Tissues Of Breast Cancer Patients Research Articles

MiRNA-195 expression in the tumor tissues of female Brazilian breast cancer patients with operable disease

OBJECTIVE:This study aimed to assess miRNA-195 expression in the tumor tissues from a cohort of Brazilian female breast cancer patients undergoing neoadjuvant chemotherapy (NAC) and evaluate its correlation with various clinicopathological markers.METHODS:Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was used to evaluate the miRNA-195 expression in tumor tissues from a cohort of female breast cancer patients undergoing NAC. This expression was then correlated with the occurrence of several distinct breast cancer molecular subtypes and other clinicopathological variables.RESULTS:A total of 55 patients were included in this study, 28 (50.9%) of whom were treated using NAC. Tumor miRNA-195 expression was suppressed in breast cancer patients, regardless of their exposure to systemic treatments, histological grade, size, nodal status, and tumor-node-metastasis (TNM) staging. This was more pronounced in luminal and triple-negative patients, and patient’s response to NAC was correlated with an increase in miRNA-195 expression.CONCLUSION:miRNA-195 is downregulated in the tumor tissues of Brazilian breast cancer patients regardless of NAC exposure; this reinforces its role as a tumor suppressor and a potential biomarker for chemotherapy response.

Lactate dehydrogenase B regulates macrophage metabolism in the tumor microenvironment.

Background: Glucose metabolism in the tumor-microenvironment is a fundamental hallmark for tumor growth and intervention therein remains an attractive option for anti-tumor therapy. Whether tumor-derived factors such as microRNAs (miRs) regulate glucose metabolism in stromal cells, especially in tumor-associated macrophages (TAMs), to hijack them for trophic support, remains elusive.Methods: Ago-RIP-Seq identified macrophage lactate dehydrogenase B (LDHB) as a target of tumor-derived miR-375 in both 2D/3D cocultures and in murine TAMs from a xenograft mouse model. The prognostic value was analyzed by ISH and multiplex IHC of breast cancer patient tissues. Functional consequences of the miR-375-LDHB axis in TAMs were investigated upon mimic/antagomir treatment by live metabolic flux assays, GC/MS, qPCR, Western blot, lentiviral knockdown and FACS. The therapeutic potential of a combinatorial miR-375-decoy/simvastatin treatment was validated by live cell imaging.Results: Macrophage LDHB decreased in murine and human breast carcinoma. LDHB downregulation increase aerobic glycolysis and lactagenesis in TAMs in response to tumor-derived miR-375. Lactagenesis reduced fatty acid synthesis but activated SREBP2, which enhanced cholesterol biosynthesis in macrophages. LDHB downregulation skewed TAMs to function as a lactate and sterol/oxysterol source for the proliferation of tumor cells. Restoring of LDHB expression potentiated inhibitory effects of simvastatin on tumor cell proliferation.Conclusion: Our findings identified a crucial role of LDHB in macrophages and established tumor-derived miR-375 as a novel regulator of macrophage metabolism in breast cancer, which might pave the way for strategies of combinatorial cancer cell/stroma cell interventions.

Collagen Type X Alpha 1 (COL10A1) Contributes to Cell Proliferation, Migration, and Invasion by Targeting Prolyl 4-Hydroxylase Beta Polypeptide (P4HB) in Breast Cancer.

BackgroundBreast cancer, a common malignant tumor, has been considered as the leading cause of cancer-related death in women. Collagen type X alpha 1 (COL10A1) is overexpressed in breast cancer. The current study was designed to determine the functional involvement and regulatory mechanism of COL10A1 on the growth and metastasis of breast cancer.Material/MethodsCOL10A1 and Prolyl 4-hydroxylase beta polypeptide (P4HB) expressions in normal tissues and tumor tissues of breast cancer patients were obtained from the GEPIA dataset. COL10A1 and P4HB levels in breast cancer cell lines were detected by real-time quantitative polymerase chain reaction (RT-qPCR) and western blot analysis. Furthermore, the interaction between COL10A1 and P4HB was confirmed by co-immunoprecipitation (Co-IP) assay. Cell Counting Kit-8 (CCK-8) and colony formation assay were applied to evaluate cell proliferation and clone-forming abilities of breast cancer cells. In addition, wound healing assay and transwell assay were performed to measure cell migration and invasion capabilities, respectively, in breast cancer.ResultsThe GEPIA dataset presented overexpressed COL10A1 and P4HB in tumor tissues of breast cancer patients. COL10A1 and P4HB expression levels were greatly upregulated in breast cancer cell lines. In addition, COL10A1 could directly interact with P4HB. Functionally, overexpressed COL10A1 boosted the proliferation and metastasis of breast cancer cells and silenced COL10A1 impeded the progression of breast cancer. More importantly, knockdown of P4HB weakened the promoting effects of overexpressed COL10A1 on cell proliferation, migration, and invasion in breast cancer.ConclusionsCOL10A1 promotes the malignant progression of breast cancer by upregulating P4HB expression, indicating that COL10A1 functions as an oncogene in breast cancer.

Forkhead box P3 and indoleamine 2,3-dioxygenase co-expression in Pakistani triple negative breast cancer patients.

BACKGROUNDForkhead box P3 (FOXP3) is a specific marker for immunosuppressive regulatory T (T-reg) cells. T-regs and an immunosuppressive enzyme, indoleamine 2,3-dioxygenase (IDO), are associated with advanced disease in cancer. AIMTo evaluate the co-expression of FOXP3 and IDO in triple negative breast cancer (TNBC) with respect to hormone-positive breast cancer patients from Pakistan. METHODSImmunohistochemistry was performed to analyze the expression of FOXP3, IDO, estrogen receptor, progesterone receptor, and human epidermal growth factor receptor on tissues of breast cancer patients (n = 100): Hormone-positive breast cancer (n = 51) and TNBC (n = 49). A total of 100 patients were characterized as FOXP3 negative vs positive and further categorized based on low, medium, and high IDO expression score. Univariate and multivariate logistic regression models were used.RESULTSOut of 100 breast tumors, 25% expressed FOXP3 positive T-regs. A significant co-expression of FOXP3 and IDO was observed among patients with TNBC (P = 0.01) compared to those with hormone-positive breast cancer. Two variables were identified as significant independent risk factors for FOXP3 positive: IDO expression high (adjusted odds ratio (AOR) 5.90; 95% confidence interval (CI): 1.22-28.64; P = 0.03) and TNBC (AOR 2.80; 95%CI: 0.96-7.95; P = 0.05). CONCLUSIONOur data showed that FOXP3 positive cells might be associated with high expression of IDO in TNBC patients. FOXP3 and IDO co-expression may also suggest its involvement in disease, and evaluation of FOXP3 and IDO expression in TNBC patients may offer a new therapeutic option.

LINC00261/microRNA-550a-3p/SDPR axis affects the biological characteristics of breast cancer stem cells.

Long non-coding RNAs (lncRNAs) have been shown to play key roles in the pathogenesis of breast cancer (BC). The study was to explore the effect of long non-coding RNA LINC00261/microRNA (miR)-550a-3p/serum deprivation response (SDPR) axis on the biological characteristics of BC stem cells (BCSCs). BC and adjacent normal tissues of patients were collected. LINC00261, miR-550a-3p and SDPR expression in BC tissues and cell lines and CD24 and CD44 expression in BC tissues was detected. CD44+ /CD24-/low BCSCs were sorted. CD44+ /CD24-/low MDA-MB-231 and MCF-7 cells were screened and transfected with altered expression of LINC00261 or miR-550a-3p to explore their roles in cell viability, microsphere (MS) formation ability, migration and invasion of CD44+ /CD24-/low BCSCs. The targeting relationships of LINC00261, miR-550a-3p and SDPR were detected. Reduced LINC00261, decreased SDPR and elevated miR-550a-3p exhibited in BC tissues of patients and cell lines. Elevated CD44+/ CD24- were present in BC tissues. LINC00261 up-regulated SDPR expression as a sponge of miR-550a-3p. Elevated LINC00261 suppressed BC cell viability, MS formation ability, migration and invasion of CD44+ /CD24-/low BCSCs. Moreover, up-regulated miR-550a-3p reversed the inhibitive effect of elevated LINC00261 on BCSCs, and reduced SDPR reversed the promotive effect of decreased miR-550a-3p on BCSCs. The study highlights that LINC00261 can adsorb miR-550a-3p to modulate SDPR, thus inhibiting the viability and MS formation of BC cells, reducing migration and invasion of CD44+ /CD24-/low BCSCs, exerting a potential effect on therapy.

Overexpression of acid-sensing ion channel 1a (ASIC1a) promotes breast cancer cell proliferation, migration and invasion.

BackgroundThe microenvironment of various tumor tissues is acidic. Acid-sensing ion channels (ASICs) are a class of ligand-gated ion channels which are sensitive to extracellular protons and are often highly expressed in tumor tissues. Breast cancer, whose extracellular microenvironment is thought to be acidic, is the most common cancer type among females in the world.MethodsThirty breast cancer tissues and adjacent normal tissues of patients were collected from 2009 to 2015 at the Xinhua hospital affiliated to Shanghai Jiao Tong University School of Medicine. The expression of acid-sensing ion channel 1a (ASIC1a), a subtype of ASICs family, was detected by immunohistochemistry in breast cancer tissues, and the effect of ASIC1a on the physiological activity of tumor cells was analyzed in vitro and in vivo experiments.ResultsIn this study, it was found that ASIC1a is highly expressed in the tissues of breast cancer patients. In vitro experiments revealed that down-regulation of ASIC1a by its antagonist PcTx-1 or ASIC1a siRNA could significantly weaken the migration, proliferation and invasion of tumor cells. In vivo studies, down-regulation or inhibition of the ASIC1a could inhibit breast tumor growth.ConclusionsThe high expression of ASIC1a might be related to the enhanced biological activity of breast cancer cells. Whether ASIC1a is a potential therapeutic target for some types of breast cancer deserves further study.

Age-related DNA methylation in paired normal and tumour breast tissue in Chinese breast cancer patients

ABSTRACT Age-related DNA methylation is a potential mechanism contributing to breast cancer development. Studies of primarily Caucasian women have identified many CpG sites of age-related methylation in non-diseased breast tissue possibly driving cancer development over time. There is a paucity of studies involving Asian women whose ages at breast cancer onset are usually younger than Caucasians. We identified the 181 most consistent age-related methylation events in non-diseased breast tissue across published studies. Age-related methylation events were measured in adjacent normal and breast tumour tissue in an exclusively Asian population at the previously identified age-related methylation sites. Age-related methylation was found in 118 probes in adjacent normal breast tissue. Methylation of 99% of these sites was increased with age and predominantly located on CpG islands in promoter regions. To ascertain biological relevance to breast cancer, we focused on the 37 sites with overall higher methylation in tumour compared to adjacent normal samples. Some sites positively related to age, including AQP5 and CORO6, inversely correlated with gene expression. Several others have known involvement in suppression of carcinogenesis including GPC5 and SST, suggesting that perturbation of epigenetic regulation at these sites due to ageing may contribute to the progression of carcinogenesis. This study highlights an age-related methylation landscape in non-tumour tissue, consistent not just across studies, but also across different populations. We present candidate age-related methylation sites warranting further investigation as potential epigenetic drivers of breast cancer. They may serve as potential targets of site-specific demethylation intervention strategies for the prevention of age-related breast cancer.

SUPT5H Post-Transcriptional Silencing Modulates PIN1 Expression, Inhibits Tumorigenicity, and Induces Apoptosis of Human Breast Cancer Cells.

Breast cancer (BrCa) is one of the most common cancers and a highly heterogenous disease, both at the pathological and molecular levels. A common element for the progression of cancer is the presence of aberrant transcription. Targeting the misregulation of transcription may serve as a tool for cancer therapeutics. SUPT5H (Suppressor of Ty 5 homolog) is a highly conserved RNA polymerase II-associated transcription elongation and processivity factor. However, few studies have examined the relationship between SUPT5H and cancer. Yeast two-hybrid and colocalization by immunofluorescence were performed to investigate protein-protein interaction. Colony formation assay, CTG assay, and crystal violet assays were performed for cell viability, clonogenicity, and cell proliferation study. Data mining was performed for expression analysis of SUPT5H in breast cancers. Flow cytometry was performed for the assessment of cell cycle and apoptosis. The Transwell chambers were employed for the migration and invasion assays. Quantitative real-time polymerase chain reaction (qRT PCR) and Western blotting were performed to measure the mRNA and protein levels of SUPT5H and other markers related to viability, migration, cell cycle, and apoptosis. Silent mutations were generated for rescue experiments. The biological function of SUPT5H was investigated through siRNA depletion of SUPT5H mRNA in vitro. We showed that SUPT5H is upregulated in breast cancer tissue as compared with the adjacent normal tissue in breast cancer patients. In human breast cancer cells, the levels of SUPT5H and PIN1 are positively correlated with each other. Our biochemical analysis showed that PIN1 interacts with SUPT5H through WW domain, that was required to promote SUPT5H protein stability. Depletion of SUPT5H by siRNA technology reduced the tumorigenic and metastatic properties, promoted s-phase cell cycle arrest and apoptosis of MDA-MB-231 cells. Moreover, depletion of SUPT5H abrogated MAPK molecules thereby regulates the oncogenic behavior of breast cancer cells. Our findings demonstrated an essential role of SUPT5H in BrCa tumorigenicity by regulating the expression levels of genes that control proliferation, migration, cell cycle, and apoptosis of breast cancer MDA-MB-231 cells.

101P Accuracy of detecting PIK3CA mutations on circulating tumour (ct) DNA as compared to tumour tissue (TT) in breast cancer (BC) patients (pts): Results from a systematic review and meta-analysis

101P Accuracy of detecting PIK3CA mutations on circulating tumour (ct) DNA as compared to tumour tissue (TT) in breast cancer (BC) patients (pts): Results from a systematic review and meta-analysis

Abstract 278: Deciphering circular RNA axes in early-onset breast cancer using a breast epithelial risk-progression 3D culture model

Abstract One of the known circular RNAs (circRNAs) functions is to sponge microRNAs (miRNAs). Studies reported dysregulated mRNA-circRNA-miRNAs axes as biomarker signatures in breast cancer (BC). Signatures in early-onset BC have not been characterized, while its incidence increased drastically worldwide. To address this gap, we performed circRNAs microarrays and miRNA-Sequencing on 3D breast epithelial risk-progression HMT-3522 S1 (S1) culture model to identify dysregulated post-transcriptional axes. S1 (non-neoplastic) cells form fully-polarized epithelium and closely mimic normal in vivo mammary epithelial morphology, while (pre-tumorigenic) Cx43KO-S1 (S1 silenced with shRNA for gap junction Cx43) lose epithelial polarity, multi-layer and mimic tumor-initiated in vivo mammary epithelial morphology. We detected through circRNA microarray (Arraystar Human circRNA Array V2) 75 significantly upregulated and 46 significantly downregulated circRNAs (Fold Change > 2, p-value < 0.05) in 6 samples of Cx43KO-S1 versus S1 cells in 3D. We then selected 10 upregulated and 8 downregulated circRNAs for validation. Of these, hsa_circ_0007961 and hsa_circ_0081481 were most significantly upregulated while hsa_circ_0060055, hsa_circ_0005185, hsa_circ_0083442 and hsa_circ_0077755 were most significantly downregulated by RT-qPCR. We focus on “possible” sponging activity of validated circRNAs to their predicted (in silico) target miRNAs, and cross-compared their dysregulation to our validated miRNA datasets (from young patients and 3D cell line). When circRNAs are upregulated, they downregulate the function of their sponged miRNAs, hence preventing them from deregulating their target mRNAs. For upregulated hsa_circ_0007961 (originating from SPRED2 mRNA), its predicted miR-99a-3p was indeed downregulated in young Lebanese and matched U.S. BC patient tissues (and in Cx43KO-S1 versus S1 microarrays) and predicted miR-653 possessed treatment potential in BC cells. For the second upregulated hsa_circ_0081481 (originating from FBXO24 mRNA), its predicted miR-3960 was exclusively dysregulated in young Lebanese BC patients. For downregulated hsa_circ_0077755 (originating from Cx43mRNA), its predicted miR-182 was commonly upregulated in young Lebanese and matched US patients and predicted miR-203a-3p was reported as promising prognostic biomarker in triple negative BC. Additionally, all predicted miRNAs targeted by all detected circRNAs were predominantly enriched in pathways in cancer. Thus, we suggest that our 3D risk-progression culture model might help delineate post-transcriptional axes to better understand early-onset BC initiation. We reveal three validated axes: i) SPRED2/hsa_circ_0007961/miR-99a-3p or miR-653-5p, ii) FBXO24/hsa_circ_0081481/miR-3960 and iii) Cx43/hsa_circ_0077755/miR-182 or miR-203, which could serve as potential biomarker signatures in early-onset BC. Citation Format: Nataly Naser Al Deen, Nadia Atallah Lanman, Shirisha Chittiboyina, Sophie Lelièvre, Heinrich Zu Dohna, Mounir AbouHaidar, Rabih Talhouk. Deciphering circular RNA axes in early-onset breast cancer using a breast epithelial risk-progression 3D culture model [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 278.

Abstract A16: The α-arrestin ARRDC3 functions as a metastasis suppressor by regulating GPCR activation of the Hippo pathway

Abstract G protein-coupled receptors (GPCRs) are key regulators of the Hippo pathway, a well-established mediator of tumorigenesis and cancer progression. The Hippo pathway is a promising therapeutic target, yet developing inhibitors for the core pathway components has proven difficult. As critical regulators of the Hippo pathway, GPCRs represent a potential upstream therapeutic target for cancer. However, despite the success and promise of GPCRs as therapeutic targets, there are currently no FDA-approved drugs targeting GPCRs for cancer. Protease-activated receptor-1 (PAR1) is a GPCR that has been shown to activate the Hippo pathway and also promote breast cancer progression. PAR1 is overexpressed in breast cancer patient tissue biopsies and in breast carcinoma cell lines and correlates with increased rates of metastasis and poor prognosis and increased invasion and metastatic potential, respectively. One mechanism that leads to PAR1 aberrant overexpression is defective lysosomal trafficking and degradation of the receptor, leading to persistent signaling. We recently showed that arrestin domain containing protein-3 (ARRDC3), an adaptor protein for E3 ubiquitin ligases, functions as a tumor suppressor by controlling proper PAR1 trafficking and degradation in highly aggressive, triple-negative breast carcinoma cells. The dysregulation of PAR1 trafficking due to loss of ARRDC3 expression leads to persistent signaling and promotes breast cancer invasion. However, the mechanism by which ARRDC3 specifically regulates PAR1-stimulated Hippo signaling to promote breast cancer metastasis remains unknown. Here, we show that ARRDC3 regulates GPCR activation of the Hippo pathway in basal-like breast cancer, where the transcriptional coactivators YAP and TAZ play differential roles in GPCR-mediated signaling and invasive capacity. ARRDC3 re-expression in invasive breast carcinoma cells attenuates PAR1-mediated activation of the Hippo pathway through regulation of TAZ but not YAP activation. Furthermore, siRNA knockdown of TAZ inhibits PAR1-stimulated Hippo signaling and invasion, whereas YAP siRNA knockdown has no effect. Our studies suggest a crucial role for ARRDC3 and TAZ in PAR1-Hippo pathway signaling in breast carcinoma invasion and metastasis. A better understanding of the mechanisms by which the Hippo pathway is regulated by GPCRs may lead to new potential therapeutic targets for the treatment or prevention of metastatic breast cancer. Citation Format: Aleena K.S. Arakaki, Wen-An Pan, Helen Wedegaertner, JoAnn Trejo. The α-arrestin ARRDC3 functions as a metastasis suppressor by regulating GPCR activation of the Hippo pathway [abstract]. In: Proceedings of the AACR Special Conference on the Hippo Pathway: Signaling, Cancer, and Beyond; 2019 May 8-11; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(8_Suppl):Abstract nr A16.

Clinical Significance of Expression Changes and Promoter Methylation of PLA2R1 in Tissues of Breast Cancer Patients

Phospholipase A2 receptor 1 (PLA2R1) expression and its role in the initiation and progression of breast cancer are an unresolved issue. PLA2R1 was found to endorse several tumor suppressive responses, including cellular senescence and apoptosis. Previous in vitro studies demonstrated that DNA hypermethylation was highly associated with the epigenetic silencing of PLA2R1 in breast cancer cell lines. Our objective was to study the level of PLA2R1 mRNA expression and the methylation of its promoter in different histological grades and molecular subtypes of breast cancer. We performed bioinformatics analyses on available human breast cancer expression datasets to assess the PLA2R1 mRNA expression. We used qRT-PCR to evaluate the PLA2R1 mRNA expression and its promoter’s methylation in breast cancer tissue in comparison to breast fibroadenomas. Our results describe, for the first time, the expression of PLA2R1 and the methylation of its promoter in human breast cancer tissues. A significant downregulation of PLA2R1, together with hypermethylation of the promoter was detected in breast cancers of different histological grades and molecular subtypes when compared to benign breast tissues. PLA2R1 promoter hypermethylation was associated with aggressive subtypes of breast cancer. In conclusion, PLA2R1 promoter hypermethylation is a potentially useful diagnostic and prognostic biomarker and could serve as a possible therapeutic target in breast cancer.

C5aR deficiency attenuates the breast cancer development via the p38/p21 axis.

Emerging evidence has shown activation of the complement component C5 to C5a in cancer tissues and C5aR expression in breast cancer cells relates to the tumor development and poor prognosis, suggesting the involvement of complement C5a/C5aR pathway in the breast cancer pathogenesis. In this study, we found that as compared to the non-tumoral tissues, both C5aR and MAPK/p38 showed an elevated expression, but p21/p-p21 showed lower expression, in the tumoral tissues of breast cancer patients. Mice deficient in C5aR or mice treated with the C5aR antagonist exhibited attenuation of breast cancer growth and reduction in the p38/p-p38 expression, but increase in p21/p-p21 expression, in the tumor tissues. Pre-treatment of the breast cancer cells with recombinant C5a resulted in reduced p21 expression, and MAPK/p38 inhibitors prevented C5a-induced reduction in p21 expression, suggesting the involvement of the MAPK/p38 signaling pathway in the C5a/C5aR-mediated suppression of p21/p-p21 expression. These results provide evidence that breast cancer development may rely on C5a/C5aR interaction, for which MAPK/p38 pathway participate in down-regulating the p21 expression. Inhibition of C5a/C5aR pathway is expected to be helpful for the treatment of patients with breast cancer.

Development of an HPLC-MS/MS Method for the Determination of Silybin in Human Plasma, Urine and Breast Tissue.

Silybin is a flavonolignan extracted from Silybum marianum with chemopreventive activity against various cancers, including breast. This study was designed to develop an HPLC-MS/MS method for the determination of silybin in human plasma, urine and breast tissue in early breast cancer patients undergoing Siliphos® supplementation, an oral silybin-phosphatidylcholine complex. The determination of silybin was carried out by liquid–liquid extraction with methyl-tert-butyl ether (MTBE); total silybin concentration was determined by treating the samples with β–glucuronidase, while for the determination of free silybin, the hydrolytic step was omitted. Naringenin and naproxen were selected as internal standards. The detection of the analyte was carried out by mass spectrometry and by chromatography. The HPLC-MS/MS method was evaluated in terms of selectivity, linearity, limit of quantification, precision and accuracy, and carryover. The method proved to be selective, linear, precise and accurate for the determination of silybin. To the best of our knowledge, this presents the first analytical method with the capacity to quantify the major bioactive components of milk thistle in three different biological matrices with a lower limit of quantification of 0.5 ng/mL for plasma. Silybin phosphatidylcholine, taken orally, can deliver high blood concentrations of silybin, which selectively accumulates in breast tumor tissue.

Dual Hypoxia-Targeting RNAi Nanomedicine for Precision Cancer Therapy.

As a hallmark of solid tumors, hypoxia promotes tumor growth, metastasis, and therapeutic resistance by regulating the expression of hypoxia-related genes. Hypoxia also represents a tumor-specific stimulus that has been exploited for the development of bioreductive prodrugs and advanced drug delivery systems. Cell division cycle 20 (CDC20) functions as an oncogene in tumorigenesis, and we demonstrated the significant upregulation of CDC20 mRNA in the tumor vs paratumor tissues of breast cancer patients and its positive correlation with tumor hypoxia. Herein, a hypoxia-responsive nanoparticle (HRNP) was developed by self-assembly of the 2-nitroimidazole-modified polypeptide and cationic lipid-like compound for delivery of siRNA to specifically target CDC20, a hypoxia-related protumorigenic gene, in breast cancer therapy. The delivery of siCDC20 by HRNPs sufficiently silenced the expression of CDC20 and exhibited potent antitumor efficacy. We expect that this strategy of targeting hypoxia-correlated protumorigenic genes by hypoxia-responsive RNAi nanoparticles may provide a promising approach in cancer therapy.

Prospecting for Breast Cancer Blood Biomarkers: Death-Associated Protein Kinase 1 (DAPK1) as a Potential Candidate.

Background Breast cancer is the commonest malignancy in women worldwide. It is estimated to affect approximately 1.5 million women annually and responsible for the greatest number of cancer-related mortalities among women. In 2018, breast cancer mortalities stood at 627,000 women representing approximately 15% of all cancer deaths among women. In Ghana, breast cancer is the second leading cause of cancer deaths, with an incidence of 2,900 cases annually; one of eight women with the disease die. This gives impetus to the fight for improved early detection, treatment, and/management. In this light, we investigated the potential of death-associated protein kinase 1 (DAPK1) as a biomarker for breast cancer. As a tumour suppressor, its expression is activated by several carcinogens to influence cellular pathways that result in apoptosis, autophagy, immune response, and proliferation. Aim To investigate DAPK1 as a blood biomarker for breast cancer. Methods Blood samples of participants diagnosed with breast cancer and healthy controls were collected and processed to obtain serum. Information on age, treatment, diagnosis, and pathology numbers was retrieved from folders. Pathology numbers were used to retrieve breast tissue blocks of patients at the Department of Pathology of the KBTH. Tissue blocks were sectioned and immunohistochemically stained with anti-DAPK1 and counterstained with hematoxylin to determine the DAPK1 expression levels. DAKP1 levels in blood sera were quantified using a commercial anti-DAPK1 ELISA kit. Case and control group means were compared using one-way ANOVA and Chi-square test. Statistical significance was set at p ≤ 0.05. Results and Discussion. DAPK1 levels were higher in sera and breast tissues of breast cancer patients than controls. The augmented DAPK1 expression can be interpreted as a stress response survival mechanism to remediate ongoing deleterious events in the cells orchestrated by carcinogenesis. In the presence of abundant DAPK1, the proliferative power of cells (both cancerous and noncancerous) is increased. This may explain why high DAPK1 expression strongly associates with aggressive breast cancer phenotypes like the ER-negative breast cancers, especially the triple-negative breast cancers (TNBC) which are the most aggressive, fast-growing, and highly metastatic. Conclusion DAPK1 is highly expressed in sera and breast tissues of breast cancer patients than nonbreast cancer participants. The elevated expression of DAKP1 in circulation rather than in breast tissues makes it a candidate for use as a blood biomarker and potential use as therapeutic target in drug development.

MiR-622 is a novel potential biomarker of breast carcinoma and impairs motility of breast cancer cells through targeting NUAK1 kinase

BackgroundAberrant expression of microRNAs (miR) has been proposed as non-invasive biomarkers for breast cancers. The aim of this study was to analyse the miR-622 level in the plasma and in tissues of breast cancer patients and to explore the role of miR-622 and its target, the NUAK1 kinase, in this context.MethodsmiR-622 expression was analysed in plasma and in tissues samples of breast cancer patients by q-RT-PCR. Bioinformatics programs, luciferase assay, public dataset analysis and functional experiments were used to uncover the role of miR-622 and its target in breast cancer cells.ResultsmiR-622 is downregulated in plasma and in tissues of breast cancer patients respect to healthy controls and its downregulation is significantly associated with advanced grade and high Ki67 level. Modulation of miR-622 affects the motility phenotype of breast cancer cells. NUAK1 kinase is a functional target of miR-622, it is associated with poor clinical outcomes of breast cancer patients and is inversely correlated with miR-622 level.ConclusionsmiR-622/NUAK1 axis is deregulated in breast cancer patients and affects the motility phenotype of breast cancer cells. Importantly, miR-622 and NUAK1 hold promises as biomarkers and as targets for breast cancers.

Mesenchymal stromal cells in adipose tissue of breast cancer patients: a cross-sectional study

Mesenchymal stromal cells (MSCs) are important components of the tumor microenvironment and are believed to facilitate cancer growth affecting cancer cells. We investigated this issue in breast cancer patients. In this cross-sectional study biopsy or mastectomy specimens from breast cancer patients who underwent surgery were evaluated based on the presence of MSCs. Flow cytometric analysis was used to evaluate the expression of CD73, CD90, CD105, and absence of CD11b or CD14, CD79, CD45, and HLA-DR expression as the criteria of MSC presence. Patients were divided into MSC (+) or (-). SPSS software was used to evaluate the association of age, tumor type, tumor size, disease grade, stage, and metastasis with the presence of MSC cells in adipose tissue of specimens. 82 patients with a mean age of 44.43±9.2 years were included in the study. MSC (+) happened in 13 (15.83) patient's samples. A regression model was conducted to evaluate the association of patient characteristics and the MSC positivity. The patient's age was associated with MSC positivity (B=-0.194, p=0.006); and the disease stage was associated with MSC positivity (B=-0.734, p=0.019). Significantly lower age of patients was seen in MSC (+) patients vs. MSC (-) patients (p=0.002). While no difference in the case of tumor type, tumor size, disease grade, stage, and metastasis was seen between two groups (P>0.05). Our result indicated that Mesenchymal stromal cells are seen more in the tumor microenvironment of younger breast cancer patients. Also, patients having mesenchymal stromal cells may have milder disease and lower disease stage.

An investigation of the relationship between TMPRSS6 gene expression, genetic variants and clinical findings in breast cancer.

Breast cancer is one of the most common types of cancer among women worldwide. The TMPRSS6 (Transmembrane Serine Protease 6) gene encodes matriptase-2, which plays an important role in iron hemostasis as the hepcidin regulator and may play a role in breast cancer susceptibility. In this study, we examined the expression levels of the TMPRSS6 gene in healthy tissues and tumor tissues of breast cancer patients; and the relationship between these levels and pathological findings. The relationship between TMPRSS6 polymorphisms (rs733655, rs5756506, rs2413450, rs855791, rs2235324, rs4820268) and patients' hematological parameters. The gene expression study encompassed 47 breast cancer patients and the gene polymorphism study consisted of 181 breast cancer patients and 100 healthy controls. Gene expression analysis was performed by qRT-PCR. The genotyping of TMPRSS6 polymorphisms was performed by RT-PCR. TMPRSS6 gene expression levels in tumor tissues were found to be 1.88 times higher than the expression levels in the control tissues. We examined the relationship between TMPRSS6 gene expression levels and pathological data, statistically significant relationship was found between patient's estrogen receptor (ER) and HER2 findings and TMPRSS6 gene expression (respectively p = 0.02, p = 0.002). When the relationship between TMPRSS6 gene polymorphisms related genotypes distributions and hematological findings was investigated, a significant relationship was identified between mean corpuscular hemoglobin concentration (MCHC) parameter and the polymorphism of only the rs733655. According to our findings, the increase in TMPRSS6 gene expression in cancerous tissues shows that matriptase-2 may be effective in the cancer process. Thus TMPRSS6 gene polymorphisms may affect the disease process by affecting the blood parameters of patients.

SH3BGRL confers innate drug resistance in breast cancer by stabilizing HER2 activation on cell membrane

BackgroundHER2-positive breast cancer is usually associated to the more aggressive progression and the worse prognosis, but the mechanism underlying the innate resistance to HER2-targeted therapy remains elusive. The scaffold protein SH3-domain-binding glutamic acid-rich protein-like protein (SH3BGRL) is indicated as a tumor suppressor in some cancers, but it is highly expressed in breast cancers. Here we characterized the tumorigenic function of SH3BGRL in HER2-expressing breast cancer cells and the subsequent effect in HER2-targeted therapies.MethodsThe interaction of SH3BGRL to HER2 were characterized with various truncated SH3BGRL mutants by immunoprecipitation and molecule docking simulation. The physiological roles of SH3BGRL interacting with HER2 in tumor progression and therapy implication were characterized by gain and loss of function approaches in vitro and in vivo. Immunohistochemistry was used for detections of SH3BGRL and p-HER2 (Y1196) expressions in xenografted tumors and human breast cancer tissues. Clinical relevance of SH3BGRL expression with HER2 was validated with both breast patient sample and the public data analyses.ResultsOur results demonstrated that SH3BGRL directly binds with HER2 on cell membrane via its motifs α1, α2 helixes and β3 sheet, which postpones HER2 internalization upon EGF stimulation. Consequently, the association between SH3BGRL and HER2 contributed to the prolonged HER2 phosphorylation at specific tyrosine sites, especially at Y1196, and their downstream signaling activation. The relevance between SH3BGRL expression and p-HER2 (Y1196) phosphorylation was validated in both xenografted tumors and the breast cancer patient tissues. Mechanistically, SH3BGRL promoted breast tumor cell proliferation and survival, while reduced the cell sensitivity to anti-tumor drugs, especially to the HER2-targeted drugs. In contrast, Silencing SH3BGRL or inhibiting its downstream signals efficiently induced apoptosis of breast tumor cells with HER2 and SH3BGRL doubly positive expression. Database analysis also highlighted that SH3BGRL is a poor prognostic marker, especially for HER2-positive breast cancers.ConclusionsOur results disclose SH3BGRL as a novel posttranslational modulator of HER2 hyperactivation, which can lead to the intrinsic resistance to HER2-targeted therapy. SH3BGRL would be a pivotal therapy target and a diagnostic marker to HER2-positve patients. Thus, targeting SH3BGRL or the downstream signaling could relieve the innate resistance to some HER2-tageted therapies for both HER2 and SH3BGRL-postive breast cancers.