Malignant Breast Tissue Research Articles

Optimized immunohistochemical detection of estrogen receptor beta using two validated monoclonal antibodies confirms its expression in normal and malignant breast tissues.

Significant controversy exists regarding the expression patterns of estrogen receptor beta (ERβ) in normal and diseased breast tissue. To address this issue, we have validated two ERβ antibodies, optimized the IHC protocols for both antibodies and now report the expression patterns of ERβ in normal and malignant breast tissues. ERβ antibody specificity was determined using western blot and IHC analysis. ERβ protein expression patterns were assessed via IHC in normal breast tissue and invasive breast carcinoma. Further, we report the detailed protocol of the ERβ IHC assay developed in our CAP/CLIA certified laboratory to provide a standardized method for future studies. We have confirmed the specificity of two independent ERβ monoclonal antibodies, one that detects total (i.e., full length plus splice variants 2-5, which do not include the ligand binding domain) ERβ protein (PPZ0506) and one that detects only the full-length form, which includes the ligand binding domain, of ERβ (PPG5/10). Using these two antibodies, we demonstrate that ERβ is highly expressed in normal human breast tissue as well as in 20-30% of invasive breast cancers. Further, these two antibodies exhibited similar staining patterns across multiple different tissues and were highly concordant with regard to determining ERβ positivity in breast cancers. ERβ protein was shown to be abundant in the majority of normal breast epithelial cells and is present in 20-30% of breast cancers. Use of these two antibodies, along with their standardized IHC protocols, provide a reference for future studies aimed at determining the utility of ERβ as a prognostic and/or predictive biomarker in various tissues of benign or malignant states.

Absence of Human Papillomavirus in Benign and Malignant Breast Tissue.

Background & Objective:Malignant breast tumors, which are one of the most important deadly cancers in women, like many other cancers, are proposed to be related to viruses etiologically. Proper management of breast carcinoma necessitates an identification of the etiological factors. Human Papillomavirus is considered to have an etiological role in breast carcinoma. We carried out this study to find out if Human Papillomavirus-DNA is present in the malignant and benign breast tissue in our patients. Methods:Seventy-five paraffin-embedded breast cancer tissues and 75 normal breast tissues and benign breast lesions were examined in this study (case-control) to look for Human Papillomavirus-DNA employing Nested Polymerase Chain reaction. The tissues were examined over a period of ten years in the pathology department of the Pathobiology Laboratory Center of Tehran.Results:No Human Papillomavirus-DNA was found in any of the malignant or control group specimens.Conclusion:Our results showed no evidence of Human Papillomavirus in cancerous and benign tissues, which is consistent with some other studies in English medical literature. More investigations using more specimens from different parts of the country are required to confirm the presence or absence of any connection between Human Papillomavirus and development of breast carcinoma in Iran.

Histopathological study of Human Papillomavirus (HPV) in breast cancer patients

A large number of epidemiological studies have evaluated the association between Human papillomavirus infection and breast carcinoma, but the results have been inconsistent.Numerous types of cytopathic effect are distinct in living cultures, but fixation and staining of the cells is essential to realize appearances of viral infection such as inclusion bodies. Identifying cytopathic effect and using it as a investigative tool needs much practice in observing stained section of numerous cell types. The present study aimed at diagnosing the histological changes in the malignant breast tissue, which caused by the HPV and comparing them with malignant breast tissue caused by other factors in order to find a way to diagnose the virus through the histopathic effect. A total of 100 samples was included in the study, 20 from benign tumor and 80 carcinomas, including different histopathological change. HPV was detected by using the conventional PCR. The breast cancer sections were examined for the expression of the effect of Human papillomavirus. In separate series the results of using formalin-fixed, paraffin -embedded sections were compared. Overall prevalence of HPV in malignant breast tissue was difference, we found that the histopathological characteristics of HPVassociated breast cancer are very different to breast cancercaused by other factors. Specifically, we showed the presence of HPV in breast cancer associates with the adenocarcinoma, The virus caused more activity in the tumor. Tumor is acinar carcinoma with increased cellularty and proliferative, While, in none infected with virus, there is no increased in cellularty or acinar, in some arease there was dilution of ducts.The observations indicate a likely causal role of HPV in causing a different histopathological change than breast cancercaused by other factors, This helps in the diagnosing of the HPV through cytopathic effect.

Docosahexaenoic Acid (DHA) Inhibits Bone Morphogenetic Protein-2 (BMP-2) Elevated Osteoblast Potential of Metastatic Breast Cancer (MDA-MB-231) Cells in Mammary Microcalcification

Microcalcification seems to be an assurance signature for the prediction of breast cancer malignancy. However, neither systematic study for deciphering the molecular mechanism of mammary microcalcification has yet been conducted, nor a mechanistic study has been performed to find out its prevention. Thus, this study firstly aimed at determining if malignant breast tissues/metastatic breast cancer cells exhibit elevated intrinsic osteoblast-like potential responsible for driving the pathological microcalcification in breast tumors. Here, tumor sample analysis showed higher levels of various osteogenic genes (e.g., Runx2, osterix), and increased ALP activity and calcification in malignant breast tissues when compared to benign tissues, indicating the existence of elevated osteoblast-like potential in malignant breast tissues as compared to benign tissues. Similarly, cell culture study found that metastatic MDA-MB-231 cells acquired a higher osteoblast-like potential as compared to less metastatic breast cancer MCF-7 cells. It was also noticed that osteoinducer bone morphogenetic protein 2 (BMP-2) increased osteoblast-like differentiation and calcification potential in breast cancer cells. Moreover, omega-3 fatty acid docosahexaenoic acid (DHA) showed an inhibitory effect on BMP-2 induced osteoblast-like potential presumably by abrogating BMP signaling. Thus, this study for the first time unraveled that DHA may mitigate microcalcification by blocking osteoblast-like potential of breast cancer cells.

A targeted mass spectrometry immunoassay to quantify osteopontin in fresh-frozen breast tumors and adjacent normal breast tissues

Osteopontin (OPN) is a multifunctional protein that can activate cell-signaling pathways and lead to cancer development and metastasis. Elevated OPN expression was reported in different cancer types, including breast tumors. Here, we present a new immuno-mass spectrometry method for OPN quantification in fresh-frozen malignant and adjacent normal human breast tissues. For quantification we used two proteotypic peptides: OPN-peptide-1 and OPN-peptide-2. Peptide concentrations were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in multiple reaction monitoring (MRM) mode with stable isotope standards (SIS) and immuno-affinity enrichment for isolation of OPN peptides. Based on the OPN-peptide-1, the average OPN concentration in normal breast tissue was 19.42 μg/g, while the corresponding level in breast tumors was 603.9 μg/g. Based on OPN-peptide-2, the average concentration in normal breast tissue was 19.30 μg/g and in breast tumors 535.0 μg/g. In ER/PR/HER2(−) patients the OPN levels in breast tumors were significantly higher than in corresponding normal breast tissue samples, whereas in the single ER/PR/HER2(+) patient the OPN concentration in tumor samples was lower than in normal breast tissue sample. In conclusion, the current method is considered promising for the quantification of OPN in research and in clinical settings and should be further studied in breast cancer patients. SignificanceA new immuno-mass spectrometry method was successfully developed and applied to determine OPN concentrations in malignant tumor and normal breast tissues from six patients, and the method is promising for OPN quantification in both research and clinical settings.

High-Risk Human Papillomaviruses and Breast Cancer in Thi-Qar province/South Iraq

The breast cancer is considered one of the most common types of cancer in Iraq, and in Thi-Qar the infections are increasing annually, it constitutes one-third of all cancer types. The HPVs are highly contagious viruses. The important aspect about HPV infections is their relationship with breast cancer. The HPV has a well-known relationship with other neoplasms in other anatomic sites and may have a role in the development of breast cancer. The HPV 16 and 18, are high-risk oncogenic viruses, and they are the most common types that can be detected in malignant tumors. To confirm the role of papillomavirus in the occurrence of breast cancer and because the absence of a study to identify the HPV genotypes in Thi-Qar province, that is necessary to determine the type of vaccine in the future, therefore, this study was conducted. The conventional PCR technique was implemented to detect the presence of HPV DNA in malignant and benign breast tissue, and the real-time PCR used to determinant HPV genotypes. This study was conducted on 100 formalin-fixed paraffin-embedded divided to 80 samples with malignant breast tumors and 20 samples of benign breast tumor were considered as a control group. The results showed that 37 (37%) of samples divided as: 32 (40%) from malignant tumors, and 5 (25%) from benign tumors gave positive results for HPV with significant differences (p<0.05), and the genotype results showed, that the HPV16 is the dominant high-risk genotype in malignant and benign tumors by positive percentage as 35.

Experience and impacts of remote telepathology consultation in improving cancer patients’ diagnosis and management in China.

e18337 Background: Over the past 4 years, we have conducted over 30 telepathology consultation cases from China covering a wide spectrum of cancers. Through our experience, we realized that the diagnostic accuracy and report integrity from Chinese pathologists is lower than US standards. which brings great concern about diagnosis, treatment and prognosis of cancer patients in China. Methods: Consultation cases were submitted by Chinese patients, their family members or physicians. Specifically, all clinical reports were sent for consultation including imaging studies, laboratory tests, pathology reports, and histology slides for review which covered a wide spectrum of malignant neoplasms, including lung, esophagus, stomach, colon, ovary, breast, prostate, neuroendocrine, melanoma, head/neck, and soft tissue sarcoma; for most cases (27/30) additional immunostaining or molecular tests were performed. All cases and reports were analyzed at top pathology institutions in the United States, including Memorial Sloan Kettering Cancer Center, Johns Hopkins Hospital, or Brigham Women’s Hospital/Beth Israel Medical Center. Following the review of the original materials, final diagnostic reports between the Chinese and US medical centers were compared for major discrepancies in diagnosis. Results: All consulted cases had major discrepancies in diagnosis (100%). Specifically, 75% of cases had improper cancer classifications or grading, 2 cases had improper quality control in immunostains rendering interpretation questionable; 3 cases lacked appropriate molecular tests; finally, 29/30 (97%) of cases lacked cancer staging based on CAP/AJCC/UICC TNM classification of cancer report protocols. Conclusions: Our consultation experiences demonstrated great discrepancies in malignant neoplasm diagnosis and reports among pathologists between the US and China. These findings raise concerns for Chinese patient’s clinical management and prognoses; moreover, they might help explain the observed difference in cancer patient's total 5-year survival between China and USA (40% vs 70%). These findings emphasize the importance of providing support and consultations for pathologists in China, especially those who practice in non-metropolitan, small county or rural local hospitals which lack updated information, technical and academic supports.

Investigation into breast cancer and partial breast reconstruction: A review.

Growing increasingly in South America, Africa and Asia, breast cancer is known as the dominant type of cancer in women. Different treatments are available for breast cancer, among which surgery is the most widely used, but researchers are trying to develop new strategies. One of the most prominent surgical methods is referred to as oncoplastic surgery, that helps to remove segments of malignant breast tissue. This type of surgery aims to obtain vast surgical margins, while the remaining tissue is rearranged so that the better cosmetic outcome is obtained. This review will investigate the breast cancer and then discuss partial breast reconstruction. Before outlining the procedures, the different types of partial breast reconstruction will be discussed. Finally, advantages and disadvantages will be outlined. MEDLINE database was used to conduct the search. The main terms used were ‘Conservation Breast Surgery Reconstruction’ AND ‘Oncoplastic Surgery’, ‘Partial Mastectomy Reconstruction’ AND ‘Conservative Breast Surgery Reconstruction’, ‘oncoplastic’ [All Fields], ‘breast’ AND ‘surgery’ OR ‘surgery’ operative’, ‘oncoplastic’ (‘breast’)’. The bibliographies of relevant papers were manually searched up to October 2018, but more recent voices are also included.

Mitochondrial autoimmunity and MNRR1 in breast carcinogenesis

BackgroundAutoantibodies function as markers of tumorigenesis and have been proposed to enhance early detection of malignancies. We recently reported, using immunoscreening of a T7 complementary DNA (cDNA) library of breast cancer (BC) proteins with sera from patients with BC, the presence of autoantibodies targeting several mitochondrial DNA (mtDNA)-encoded subunits of the electron transport chain (ETC) in complexes I, IV, and V.MethodsIn this study, we have characterized the role of Mitochondrial-Nuclear Retrograde Regulator 1 (MNRR1, also known as CHCHD2), identified on immunoscreening, in breast carcinogenesis. We assessed the protein as well as transcript levels of MNRR1 in BC tissues and in derived cell lines representing tumors of graded aggressiveness. Mitochondrial function was also assayed and correlated with the levels of MNRR1. We studied the invasiveness of BC derived cells and the effect of MNRR1 levels on expression of genes associated with cell proliferation and migration such as Rictor and PGC-1α. Finally, we manipulated levels of MNRR1 to assess its effect on mitochondria and on some properties linked to a metastatic phenotype.ResultsWe identified a nuclear DNA (nDNA)-encoded mitochondrial protein, MNRR1, that was significantly associated with the diagnosis of invasive ductal carcinoma (IDC) of the breast by autoantigen microarray analysis. In focusing on the mechanism of action of MNRR1 we found that its level was nearly twice as high in malignant versus benign breast tissue and up to 18 times as high in BC cell lines compared to MCF10A control cells, suggesting a relationship to aggressive potential. Furthermore, MNRR1 affected levels of multiple genes previously associated with cancer metastasis.ConclusionsMNRR1 regulates multiple genes that function in cell migration and cancer metastasis and is higher in cell lines derived from aggressive tumors. Since MNRR1 was identified as an autoantigen in breast carcinogenesis, the present data support our proposal that both mitochondrial autoimmunity and MNRR1 activity in particular are involved in breast carcinogenesis. Virtually all other nuclear encoded genes identified on immunoscreening of invasive BC harbor an MNRR1 binding site in their promoters, thereby placing MNRR1 upstream and potentially making it a novel marker for BC metastasis.

Towards mm-wave spectroscopy for dielectric characterization of breast surgical margins

PurposeThe evaluation of the surgical margin in breast conservative surgery is a matter of general interest as such treatments are subject to the critical issue of margin status as positive surgical margins can undermine the effectiveness of the procedure. The relatively unexplored ability of millimeter-wave (mm-wave) spectroscopy to provide insight into the dielectric properties of breast tissues was investigated as a precursor to their possible use in assessment of surgical margins. MethodsWe assessed the ability of a mm-wave system with a roughly hemispherical sensitive volume of ∼3 mm radius to distinguish malignant breast lesions in prospectively and consecutively collected tumoral and non-tumoral ex-vivo breast tissue samples from 91 patients. We characterized the dielectric properties of 346 sites in these samples, encompassing malignant, fibrocystic disease and normal breast tissues. An expert pathologist subsequently evaluated all measurement sites. ResultsAt multivariate analysis, mm-wave dielectric properties were significantly correlated to histologic diagnosis and fat content. Further, using 5-fold cross-validation in a Bayesian logistic mixed model that considered the patient as a random effect, the mm-wave dielectric properties of neoplastic tissues were significantly different from normal breast tissues, but not from fibrocystic tissue. ConclusionReliable discrimination of malignant from normal, fat-rich breast tissue to a depth compatible with surgical margin assessment requirements was achieved with mm-wave spectroscopy.

Abstract P3-09-02: CRISPR-Cas9 mediated BRCA1 mutation in primary cells: Mutation efficiency and effects

Abstract Background: Germline mutations in Breast Cancer Associated (BRCA) 1 or 2 genes confer an increased risk of the development of breast and ovarian cancer. Germline mutation is followed by somatic loss of heterozygosity (LOH) resulting in biallelic inactivation. BRCA1 is involved in multiple homeostatic functions including control of chromatin organization, gene transcription, protein stability and cell division. Recent studies have demonstrated heterogeneity in LOH within and between premalignant and malignant breast tissues of BRCA1 mutation carriers. We hypothesize that LOH does not have a unitary effect on phenotype but differs by the function that is abrogated. Methods: To test our hypothesis, we adopted CRISPR-Cas9 gene editing technology. The guide RNAs for targeting the exon sequence in the RING finger, nuclear export signal (NES), nuclear localization signal (NLS) and BRCA1 C Terminus (BRCT) domain/motif of BRCA1 were designed and synthesized. MCF10A cells were transfected with a complex of guide RNA and Cas9 protein (RNP) to cause in/del mutation. The mutation was analyzed by both T7E1 assay, and an innovative and more precise method developed in our lab that utilizes linked nucleic acids (LNA) and qPCR. Proliferation and apoptosis assays were performed using the transfected cells. Organoids prepared from BRCA1 mutation carriers also were transfected with RNPs and the mutation burden determined. Results: Since single cell clones of the transfected MCF10A cells could not be selected and expanded, a pool of transfected cells was used for the analyses. T7E1 assay and qPCR analysis using LNAs demonstrated the presence of the mutations. A standard curve was created to enable the calculation of the mutation burden. IncuCyte analysis revealed increased proliferation and apoptosis, induced by irradiation, in cells with the mutation in Exon 10, where the extent of increase varied from 11% to 48% depending on the degree of mutation. In contrast, cells with the mutation in Exon 5 displayed diminished proliferation with no change in apoptosis. That mutations in exon10 and 5 have distinct biological effects when compared to the mutations in other exons is intriguing, and modification of binding proteins will be investigated. Organoids generated from BRCA1 mutation carriers (primary and nonmalignant cells) were able to be successfully transfected using the NEON electroporation system. Mutations were introduced by the CRISPR-Cas9 system and their extent quantified by our LNA-mediated qPCR method. Conclusions: CRISPR-Cas9-mediated gene editing of BRCA1 in MCF10A resulted in a change in the proliferation rate and the extent of apoptosis that is dependent on the location of the de novo mutation within the gene. The development of a novel method, LNA-mediated qPCR, provides quantitative information regarding the mutations that may be used to correlate mutation burden with biological functional change. Successful establishment of this BRCA1 tumorigenesis model has provided us with a method to test other putative tumor suppressors. Citation Format: Choi MR, Yadav S, Shidfar A, Khan SA, Clare SE. CRISPR-Cas9 mediated BRCA1 mutation in primary cells: Mutation efficiency and effects [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-09-02.

Quantitative label-free mass spectrometry using contralateral and adjacent breast tissues reveal differentially expressed proteins and their predicted impacts on pathways and cellular functions in breast cancer

Proteins play an essential role in the biological processes associated with cancer. Their altered expression levels can deregulate critical cellular pathways and interactive networks. In this study, the mass spectrometry-based label-free quantification followed by functional annotation was performed to investigate the most significant deregulated proteins among tissues of primary breast tumor (PT) and axillary metastatic lymph node (LN) and corresponding non-tumor tissues contralateral (NCT) and adjacent (ANT) from patients diagnosed with invasive ductal carcinoma. A total of 462 proteins was observed as differentially expressed (DEPs) among the groups analyzed. A high level of similarity was observed in the proteome profile of both non-tumor breast tissues and DEPs (n = 12) were mainly predicted in the RNA metabolism. The DEPs among the malignant and non-tumor breast tissues [n = 396 (PTxNCT) and n = 410 (LNxNCT)] were related to pathways of the LXR/RXR, NO, eNOS, eIF2 and sirtuins, tumor-related functions, fatty acid metabolism and oxidative stress. Remarkable similarity was observed between both malignant tissues, which the DEPs were related to metastatic capabilities. Altogether, our findings revealed differential proteomic profiles that affected cancer associated and interconnected signaling processes. Validation studies are recommended to demonstrate the potential of individual proteins and/or pathways as biological markers in breast cancer. SignificanceThe proteomic analysis of this study revealed high similarity in the proteomic profile of the contralateral and adjacent non-tumor breast tissues. Significant differences were identified among the proteome of the malignant and non-tumor tissue groups of the same patients, providing relevant insights into the hallmarks, signaling pathways, biological functions, and interactive protein networks that act during tumorigenesis and breast cancer progression. These proteins are suggested as targets of relevant interest to be explored as potential biological markers related to tumor development and metastatic progression in the breast cancer disease.

The role of miRNAs as a predictor of multicentricity in breast cancer.

Expression profiles of miRNAs are shown to be different in various cancers to regulate expression of mRNA or to have a role in inhibition of translation, thus it shows the possible effect in progression, invasion and metastasis of breast cancer cells. The effect of breast conserving treatment in local recurrence and survival rates for the patients who have multicentric breast cancer is still controversial. In our study, we intended to evaluate the foresight of 84 miRNAs which are identified in breast cancer for having differentiated expressions. Thirty-one patients with unifocal and 26 patients with multicentric breast cancer were included in this study. These tissue samples of both malignant and normal breast tissues were kept in RNA later solution at - 80°C. Eighty-four miRNAs were studied with miScript miRNA PCR Array Human Breast Cancer kit. Fold change, cut off value was accepted as four. In unifocal group, there were 13 upregulated and five downregulated miRNAs and in multicentric group, there were three upregulated and seven downregulated miRNAs. To reach better results for breast cancer diagnosis and treatment, it is important to enlighten tumor biology, and pay attention to target and individual therapy. Thus, miRNAs have potential role in identifying tumor characteristics in supporting diagnosis and resulting with better evaluated disease for better treatment results with individual strategies.

Tissue spatial correlation as cancer marker.

.We propose an intrinsic cancer marker in fixed tissue biopsy slides, which is based on the local spatial autocorrelation length obtained from quantitative phase images. The spatial autocorrelation length in a small region of the tissue phase image is sensitive to the nanoscale cellular morphological alterations and can hence inform on carcinogenesis. Therefore, this metric can potentially be used as an intrinsic cancer marker in histopathology. Typically, these correlation length maps are calculated by computing two-dimensional Fourier transforms over image subregions—requiring long computational times. We propose a more time-efficient method of computing the correlation map and demonstrate its value for diagnosis of benign and malignant breast tissues. Our methodology is based on highly sensitive quantitative phase imaging data obtained by spatial light interference microscopy.

TRAIL and TRAIL Receptors as Prognostic Markers in Breast Cancer Patients

Background: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a key player in the extrinsic pathway of apoptosis; it selectively damages cancer cells through binding to its surface receptors, however, cancers can escape this pathway through expression of dysfunctional decoy receptors. Purpose: The present study directed mainly to elucidate the serum TRAIL levels in breast cancer patients and to explore the variation in gene expression of TRAIL death and decoy receptors in breast cancer tissues, and to explore their role as prognostic markers in breast cancer as well as to detect their correlation with Patients’ Clinical Characteristics. Subjects and Methods: TRAIL levels were assayed in the sera of 124 breast cancer patients and 150 healthy females. Moreover, the expression of TRAIL death and decoy receptors was determined in both malignant and adjacent normal breast tissues collected from patients. ER, PR and Her-2 expression in breast cancer tissue were performed using immunohistochemical method. Apoptotic index (AI) was analyzed using H&E stain under light microscopy. Results: Serum levels of TRAIL in breast cancer patients were significantly lower than controls (P < 0.001), additionally, the expression of DR4, DR5 and DcR1 were significantly up-regulated (p < 0.001, p < 0.001, p = 0.039, respectively), however, no significant difference was found in the expression of DcR2 in breast cancer tissues as compared to the corresponding normal tissues. Moreover, the apoptotic index in breast cancer tissues was significantly higher than the corresponding normal tissues. On the other side, decreased Serum TRAIL levels and increased DcR1 expression were associated with decreased overall patients’ survival. Conclusions: The expression of both DR4 and DR5 is required for TRAIL-induced apoptosis in breast cancer tissues; in addition, serum TRAIL and profiling of TRAIL receptors expression may serve as prognostic markers in breast cancer patients.

Analysis of PKC-ζ protein levels in normal and malignant breast tissue subtypes.

It is estimated that breast cancer will be the second leading cause of cancer-associated mortality in women in 2018. Previous research has demonstrated that the atypical protein kinase C-ζ (PKC-ζ) is a component of numerous dysregulated pathways in breast cancer, including cellular proliferation, survival, and cell cycle upregulation. The present study investigated the PKC-ζ protein in breast tissue to evaluate its potential as a biomarker for breast cancer invasion, and demonstrated that an overexpression of PKC-ζ protein can be indicative of carcinogenesis. The present study analyzed the expression of PKC-ζ in individuals with no tumor complications and malignant female human breast tissue samples (lobular carcinoma in situ, invasive lobular carcinoma, ductal carcinoma in situ and invasive ductal carcinoma) with the use of western blot analysis, immunohistochemistry and statistical analysis (83 samples). The present study also evaluated the invasive behavior of MDA-MB-231 breast cancer cells following the knockdown of PKC-ζ with a Transwell invasion assay and an immunofluorescent probe for filamentous actin (F-actin) organization. The data demonstrated that PKC-ζ expression was identified to be higher in invading tissues when compared with non-invading tissues. The results also suggest that PKC-ζ is more abundant in ductal tissues when compared with lobular tissues. In addition, the protein studies also suggest that PKC-ζ is a component for invasive behavior through the Ras-related C3 botulinum toxin substrate 1 (Rac1) and Ras homolog gene family member A (RhoA) pathway, and PKC-ζ is required for the F-actin reorganization in invasive cells. Therefore, PKC-ζ should be considered to be a biomarker in the development of breast cancer as well as an indicator of invading tumor cells.

Mammographic density and breast tissue expression of inflammatory markers, growth factors, and vimentin

BackgroundMammographic density is a known risk factor for breast cancer, but the underlying pathologic characteristics are not well understood. The current analysis investigated the expression of several markers of interest, e.g., inflammation and growth, with mammographic density (MD) in normal and malignant breast tissue specimens from 279 women of the Multiethnic Cohort (MEC).MethodsBreast cancer cases, recruited from a nested case-control study within the MEC, provided mammograms for density evaluation. Protein expression (COX-2, TNF-α, TGF-β, IGF-1R, IGFBP-2, and vimentin) was assessed by immunohistochemical detection. Linear regression was applied to evaluate the relation between marker expression and percent density and to compute adjusted means with 95% confidence intervals (CI) by marker status while adjusting for confounders.ResultsDue to missing cores and tissue, normal tissue could only be evaluated for COX-2 and vimentin. No significant associations with mammographic density were detected for all markers analyzed. For inflammatory markers (TNF-α, COX-2, and TGF-β) in tumor tissue, MD were non-significantly higher with stronger expression but the differences were very small. For example, the mean MD values for no, weak, and strong TNF-α expression were 35% (95% CI 24–47%), 39% (95% CI 29–48%), and 38% (95% CI 27–50%). In a posthoc analysis among postmenopausal women only, the difference across categories of TNF-α expression increased to 25% (95% CI 12–39%), 35% (95% CI 23–48%), and 35% (95% CI 20–49%).ConclusionsThe current analysis offers little support for an involvement of immunohistochemical markers representing inflammatory and growth factor pathways as predictors of breast density.

Targeting TPX2 suppresses proliferation and promotes apoptosis via repression of the PI3k/AKT/P21 signaling pathway and activation of p53 pathway in breast cancer

Targeting protein for Xenopus kinesin-like protein 2 (TPX2) is a microtubule-associated protein required for mitosis and spindle assembly. Previous studies showed that TPX2 is overexpressed in various human cancers and promotes cancer progression. In this study, the differentially expressed genes including TPX2 were screened in GEO database for gene expression microarray of breast cancer. The TPX2 expression level was significantly increased in breast cancer cells and the breast malignant tissues compared with those controls. In vitro experiment further confirmed that knockdown of TPX2 by small hairpin RNA inhibited breast cancer cell proliferatio, migration, and induced cell apoptosis. TPX2 silencing decreased the expression of PI3K and extent of AKT phosphorylation, as well as increased expression of p53 and p21. Taken together, our findings indicate that TPX2 silencing negatively regulates the PI3K/AKT and activates p53 signaling pathway by which breast cancer cells proliferation were inhibited whereas cellulars apoptosis were accelerated, suggesting that TPX2 may be a potential target for anticancer therapy in breast cancer.

Second-harmonic patterned polarization-analyzed reflection confocal microscopy of stromal collagen in benign and malignant breast tissues

We present the results of polarimetric analysis of collagen on varying pathologies of breast tissues using second-harmonic patterned polarization-analyzed reflection confocal (SPPARC) microscopy. Experiments are conducted on a breast tissue microarray having benign tissues (BT), malignant invasive lobular carcinoma (ILC), and benign stroma adjacent to the malignant tissues (called the benign adjacent tissue, or BAT). Stroma in BAT and ILC exhibit the largest parameter differences. We observe that stromal collagen readings in ILC show lower depolarization, lower diattenuation and higher linear degree-of-polarization values than stromal collagen in BAT. This suggests that the optical properties of collagen change most in the vicinity of tumors. A similar trend is also exhibited in the non-collagenous extrafibrillar matrix plus cells (EFMC) region. The three highlighted parameters show greatest sensitivity to changes in the polarization response of collagen between pathologies.

Identification of breast tumors from diathermy smoke by differential ion mobility spectrometry

IntroductionBreast cancer is the most frequent cancer in women worldwide. The primary treatment is breast-conserving surgery or mastectomy with an adequate clearance margin. Diathermy blade is used extensively in breast-conserving surgery. Surgical smoke produced as a side product has cancer-specific molecular features. Differential mobility spectrometry (DMS) is a rapid and affordable technology for analysis of complex gas mixtures. In our study we examined surgical smoke from malignant and benign breast tissue created with a diathermy blade using DMS. Material and methodsPunch biopsies of 4 mm diameter from breast cancer surgical specimens were taken during gross dissection of fresh surgical specimen and placed in a well plate. The measurement system is a custom-built device called automatic tissue analysis system (ATAS) based on a DMS sensor. Each specimen was incised with a diathermy blade and the surgical smoke was analyzed. ResultsWe examined 106 carcinoma samples from 21 malignant breast tumors. Benign samples (n = 198) included macroscopically normal mammary gland (n = 82), adipose tissue (n = 88) and vascular tissue (n = 28). The classification accuracy when comparing malignant samples to all benign samples was 87%. The sensitivity was 80% and the specificity was 90%. The classification accuracy of carcinomas to ductal and lobular was 94%, 47%, respectively. ConclusionsBenign and malignant breast tissue can be identified with ATAS. These results lay foundation for intraoperative margin assessment with DMS from surgical smoke.