Carbonic Anhydrase IX in Tumor Tissue and Plasma of Breast Cancer Patients: Reliable Biomarker of Hypoxia and Prognosis.

Carbonic anhydrase IX (CA IX) is recognized as an excellent marker of hypoxia and an adverse prognostic factor in solid tumors, including breast cancer (BC). Clinical studies confirm that soluble CA IX (sCA IX), shed into body fluids, predicts the response to some therapeutics. However, CA IX is not included in clinical practice guidelines, possibly due to a lack of validated diagnostic tools. Here, we present two novel diagnostic tools-a monoclonal antibody for CA IX detection by immunohistochemistry and an ELISA kit for the detection of sCA IX in the plasma-validated on a cohort of 100 patients with early BC. We confirm that tissue CA IX positivity (24%) correlates with tumor grading, necrosis, negative hormone receptor status, and the TNBC molecular subtype. We show that antibody IV/18 can specifically detect all subcellular forms of CA IX. Our ELISA test provides 70% sensitivity and 90% specificity. Although we showed that this test could detect exosomes in addition to shed CA IX ectodomain, we could not demonstrate a clear association of sCA IX with prognosis. Our results indicate that the amount of sCA IX depends on subcellular CA IX localization, but more strictly on the molecular composition of individual molecular subtypes of BC, particularly on metalloproteinases inhibitor expression.

Radar reflector guidance system in breast surgery: A single-institution feasibility study.

Mammography breast cancer screening programs and continuing improvements in early diagnosis of the disease have led to more frequent detection of nonpalpable breast lesions. The commonly used technique in guiding the surgical removal of these lesions is hook wire-guided localization (WGL). However, the WGL procedure has been criticized for the last years. Key disadvantages of WGL are possible wire transection, wire migration before or during surgery, patient discomfort and pneumothorax. Over the last decade, alternatives to wire localization have emerged. In this study the authors present their initial experience with a wireless, nonradioactive, wave reflection implant system that enables surgeons to safely and accurately remove breast lesions (Tab. 2, Fig. 4, Ref. 20). Keywords: breast cancer, breast surgery, nonpalpable lesions, preoperative localization.

Intraoperative evaluation of metastases in sentinel nodes through one-step nucleic acid amplification: Initial experience in a single institution.

The objective of this study was to gain our initial experience in one-step nucleic acid amplification (OSNA) for detecting sentinel lymph node (SLN) metastasis as compared to standard pathological staging in patients with breast cancer. Fifteen patients with preoperatively confirmed early breast cancer eligible for breast­saving therapy and sentinel lymph node biopsy (SLNB) were enrolled in the study. Lymphatic mapping and SLNs detection were performed through the magnetic method. Excised SLNs were intraoperatively examined through OSNA and frozen­section methods. All lymph nodes were postoperatively examined through histopathology and immunohistochemistry. The results of latter methods were correlated. Our initial experience proved OSNA to be a sensitive and efficient alternative to intraoperative assessment of metastases in SLN in breast cancer patients. Moreover, the information obtained by the OSNA method provides the surgeon with the possibility of assessing a more accurate prognosis during the initial surgery (Tab. 3, Fig. 4, Ref. 36). Text in PDF www.elis.sk Keywords: breast cancer, metastases, surgery, sentinel nodes, OSNA.

Expression of COX-2, p16, and Ki67 in the range from normal breast tissue to breast cancer.

The increasing number of diagnosed breast lesions lead to the critical need for new markers that would elucidate the process of tumorigenesis. The objective of the study was to examine COX-2, p16, and Ki67 expression in a broad spectrum of breast lesions in order to define the proteins' phenotype throughout the tumorigenesis. Expression was studied by immunohistochemistry in 308 human breast samples divided into 7 subgroups - flat epithelial atypia (FEA), atypical hyperplasia (ADH), intraductal carcinoma (DCIS), invasive cancer (IC), benign lesions (BLs), normal tissue adjacent to breast cancer (CANT), and fatty tissue (FT). Analysis among 4 subgroups - premalignant lesions (DIN), IC, BLs, and normal tissue was also performed. High prevalence of COX-2 overexpression was found in all breast lesions including BLs (70% FEA, 89% ADH, 86% DCIS, 81% IC, 44% CANT, 92% BLs, 29% FT). Significant dominance of p16 overexpression was found in premalignant lesions and BLs (50% FEA, 67% ADH, 50% DCIS, 37% IC, 8% CANT, 58% BLs, 21% FT). The location of staining within p16+ cells differed - BLs showed nuclear positivity, whereas in IC it was exclusively cytoplasmic. Premalignant lesions showed all types of p16 positivity. Significantly higher prevalence of COX-2+p16+Ki67+ phenotype was in premalignant tumors with the highest prevalence in ADH (40% of FEA, 67% ADH, 35% DCIS, 20% IC, 3% CANT, 20% BLs, 14% FT). Our observations showed a high prevalence of COX-2+p16+Ki67+ phenotype in premalignant lesions. Further studies are needed in order to elucidate if this phenotype reflects any specific pathway of future progression of premalignant breast lesions.

CAIX-Mediated Control of LIN28/let-7 Axis Contributes to Metabolic Adaptation of Breast Cancer Cells to Hypoxia.

Solid tumors, including breast cancer, are characterized by the hypoxic microenvironment, extracellular acidosis, and chemoresistance. Hypoxia marker, carbonic anhydrase IX (CAIX), is a pH regulator providing a selective survival advantage to cancer cells through intracellular neutralization while facilitating tumor invasion by extracellular acidification. The expression of CAIX in breast cancer patients is associated with poor prognosis and metastases. Importantly, CAIX-positive hypoxic tumor regions are enriched in cancer stem cells (CSCs). Here we investigated the biological effects of CA9-silencing in breast cancer cell lines. We found that CAIX-downregulation in hypoxia led to increased levels of let-7 (lethal-7) family members. Simultaneously with the increase of let-7 miRNAs in CAIX-suppressed cells, LIN28 protein levels decreased, along with downstream metabolic pathways: pyruvate dehydrogenase kinase 1 (PDK1) and phosphorylation of its substrate, pyruvate dehydrogenase (PDH) at Ser-232, causing attenuation of glycolysis. In addition to perturbed glycolysis, CAIX-knockouts, in correlation with decreased LIN28 (as CSC reprogramming factor), also exhibit reduction of the further CSC-associated markers NANOG (Homeobox protein NANOG) and ALDH1 (Aldehyde dehydrogenase isoform 1). Oppositely, overexpression of CAIX leads to the enhancement of LIN28, ALDH1, and NANOG. In conclusion, CAIX-driven regulation of the LIN28/let-7 axis augments glycolytic metabolism and enhances stem cell markers expression during CAIX-mediated adaptation to hypoxia and acidosis in carcinogenesis.

Histopathological characterization of ulcerated breast cancer and comparison to their non-ulcerated counterparts.

Approximately 6-15 % of breast cancer patients are diagnosed with primary ulcerated breast cancer (ULBC). ULBC is known to be associated with short recurrence free and poor overall survival. Therefore, the purpose of this study was to characterize ULBC and compare the histopathological findings with those of non-ulcerative breast cancer (NULBC). A total of 152 ULBCs were evaluated and compared to 304 consecutive non-ulcerated, age-matched breast malignancies. Patients mean age was 65 years (SD = 13.0 ULBC, SD = 14.0 NULBC). ULBC was associated with a higher rate of poorly differentiated tumors (p = < 0.001), as well as larger tumor sizes (p = < 0.001). As expected, the rate of axillary lymph node involvement was higher in ULBC patients (p = <0.001). In addition to that, ULBC was associated with a higher rate of triple negative breast cancer (p = 0.002), and higher Ki67 expression (p = < 0.001). ULBC showed more aggressive histopathological features in comparison to NULBC which may contribute to the generally known poorer prognosis of women with ULBC.

Mammary myofibroblastoma: Presentation of case.

INTRODUCTION AND IMPORTANCE: Mammary myofibroblastoma (MFB) is a rare benign mesenchymal lesion. It belongs to the family of benign spindle cell tumours of the mammary stroma and may exhibit confusing variants. Some of them may mimic invasive tumours, leading to the diagnostic dilemmas, especially in core needle biopsy specimens or frozen sections. Familiarity with the characteristics of this tumour is of great importance for accurate diagnosis and proper treatment. CASE PRESENTATION: We report about a rare form of CD34-negative mixed epithelioid/lipomatous form of mammary myofibroblastoma in a 48-year-old Caucasian premenopausal woman with no previous medical history. Breast imaging suggested a benign lesion. The core needle biopsy suggested breast MFB. The definitive diagnosis was established through histopathology and immunohistochemistry of the lumpectomy specimen. CLINICAL DISCUSSION: Despite its rarity, breast MFB is a disease with a wide spectrum of histologic morphologies. CD34 positivity is seen in majority of MFB cases. MFBs uncommonly show absent expression of CD34, a potential diagnostic pitfall, just like in our case. CONCLUSION: Pathologists should recognise the wide range of differential diagnoses and be familiar with the diverse morphological appearances of these lesions to make an accurate diagnosis. Surgical excision is at present the ordinary treatment of MFB.

Secretory carcinoma of the breast: A case report.

INTRODUCTION: Secretory breast carcinoma (SBC) is a rare breast tumor which accounts for < 0.15% of all breast cancers. It was originally described as a juvenile breast carcinoma, occurring in young children and adolescent women. SBC is associated with a characteristic balanced translocation, t(12;15), that creates aETV6-NTRK3 gene fusion. PRESENTATION OF CASE: A 52-year-old Caucasian woman had palpable lump in her right breast. After breast imaging examination (BI-RADS 4b) and preoperative core-needle biopsy with suspicion of SBC a breast conserving therapy was performed. The diagnosis of SBC was confirmed through immunohistochemistry and cytogenetic examination of the tumor. The patient is now 22 months post­surgery and remains disease­free. DISCUSSION: Recent studies reported that the disease occurs at a later age than previously recognized, and is associated with good long-term survival. In breast imaging it may mimic a benign tumor. Immunohistochemistry and cytogenetic analysis of the tumor are crucial for confirmation of SBC. CONCLUSION: There is no consensus with regard to the best treatment strategy for patients with SBC. Breast conserving therapy with sentinel lymph nodes biopsy is at present the first choice treatment. Further research for a specific NTRK3 tyrosine kinase inhibitor could lead to the discovery of a new targeted treatment of this tumor.

Localization of impalpable breast lesions and detection of sentinel lymph nodes through magnetic methods.

PURPOSE: The most commonly used technique for guiding the surgical removal of impalpable breast lesions is wire-guided localization (WGL). Potential complications of WGL include wire migration, wire transection, patient discomfort, and pneumothorax. Recently, another possibility for preoperative localization of breast lesions trough small steel seeds was developed. A magnetic handheld probe can be used both for localization of breast lesions and sentinel lymph nodes (SLNs) detection. METHODS: In this study, we used a new technology for localizing breast lesions in conjunction with sentinel nodes (SLNs) detection through SPIO nanoparticles; both detected using a magnetic probe. The technique uses small steel markers (Magseed®) with magnetic properties, which are placed in breasts under ultrasonographic or mammographic guidance. 41 localization seeds were placed in 38 patients. In 27 patients with malignant tumors, simultaneous use of magnetic method for SLNs detection was used. RESULTS: In all 38 patients, breast lesions were accurately localized using this method. No interference between Magseed signals and SPIO tracer signals were observed during magnetic probe measurements. All tumors were exscised with tumor-free surgical margins. The SLN biopsy was successful in all patients undergoing this procedure. The SLN median detection rate was 3 nodes. CONCLUSIONS: The new magnetic methods are reliable alternatives for localizing breast lesions and SLN detection. They are well tolerated by patients and they can avoid the disadvantages of WGL. They have the potential to make tumor localization and SLN biopsy procedures possible in facilities without a nuclear medicine department or where radioisotope availability is limited.

Genetic counselling and testing of susceptibility genes for therapeutic decision-making in breast cancer-an European consensus statement and expert recommendations.

An international panel of experts representing 17 European countries and Israel convened to discuss current needs and future developments in BRCA testing and counselling and to issue consensus recommendations. The experts agreed that, with the increasing availability of high-throughput testing platforms and the registration of poly-ADP-ribose-polymerase inhibitors, the need for genetic counselling and testing will rapidly increase in the near future. Consequently, the already existing shortage of genetic counsellors is expected to worsen and to compromise the quality of care particularly in individuals and families with suspected or proven hereditary breast or ovarian cancer. Increasing educational efforts within the breast cancer caregiver community may alleviate this limitation by enabling all involved specialities to perform genetic counselling. In the therapeutic setting, for patients with a clinical suspicion of genetic susceptibility and if the results may have an immediate impact on the therapeutic strategy, the majority voted that BRCA1/2 testing should be performed after histological diagnosis of breast cancer, regardless of oestrogen receptor and human epidermal growth factor receptor 2 (HER2) status. Experts also agreed that, in the predictive and therapeutic setting, genetic testing should be limited to individuals with a personal or family history suggestive of a BRCA1/2 pathogenic variant and should also include high-risk actionable genes beyond BRCA1/2. Of high-risk actionable genes, all pathological variants (i.e. class IV and V) should be reported; class III variants of unknown significance, should be reported provided that the current lack of clinical utility of the variant is expressly stated. Genetic counselling should always address the possibility that already tested individuals might be re-contacted in case new information on a particular variant results in a re-classification.

Effect of tibolone on breast cancer cell proliferation in postmenopausal ER+ patients: results from STEM trial.

PURPOSE: Tibolone is a selective tissue estrogenic activity regulator, approved for the treatment of vasomotor symptoms in postmenopausal women. We have done an exploratory, double-blind, randomized, placebo-controlled pilot trial to investigate the tissue-specific effects of 2.5 mg tibolone on breast cancer in postmenopausal women, in particular on tissue proliferation (STEM, Study of Tibolone Effects on Mamma carcinoma tissue). EXPERIMENTAL DESIGN: Postmenopausal women with initially stage I/II, estrogen receptor-positive (ER+) primary breast cancer, were randomly assigned to 14 days of placebo or 2.5 mg/d tibolone. Core biopsies of the primary tumor were obtained before and after treatment. Ki-67 and apoptosis index were analyzed in baseline and corresponding posttreatment specimen. RESULTS: Of 102 enrolled patients, 95 had evaluable data. Baseline characteristics were comparable between both treatment groups. Breast cancer cases are mainly invasive (99%), stage I or II (42% and 50% respectively), and ER+ (99%). Median intratumoral Ki-67 expression at baseline was 13.0% in the tibolone group and 17.8% in the placebo group, and decreased to 12.0% after 14 days of tibolone while increasing to 19.0% in the placebo group. This change from baseline was not significantly different between tibolone and placebo (Wilcoxon test; P=0.17). A significant difference was observed between the treatment groups when the median change from baseline apoptosis index was compared between the treatment groups (tibolone, 0.0%; placebo, +0.3%; Wilcoxon test; P=0.031). The incidence of adverse effects was comparable. CONCLUSIONS: In ER+ breast tumors, 2.5 mg/d tibolone given for 14 days has no significant effect on tumor cell proliferation.

Granular cell tumor in axillary region: A rare entity.

A granular cell tumor (GCT), is a rare soft tissue tumor which may occur throughout the body, usually in the head and neck, skin or subcutaneous tissues of the trunk and upper extremities, and female genital region. A total of 5-8% of all cases of GCTs occur in the breast. GCT of the breast may mimic breast cancer both clinically and radiologically. GCTs are usually benign and solitary; however, approximately 2% occur as malignant tumors. Benign GCTs are treated with wide local excision and are associated with a good prognosis. The current case report presents findings in a patient with a benign form of GCT in a rare location, specifically in the axillary region.

Simultaneous use of magnetic method in localization of impalpable breast cancer and sentinel lymph nodes detection: initial experience.

AIM: In this study we used a new technology for localization of non-palpable breast tumors using a small steel marker in conjunction of sentinel nodes (SLNs) detection through injection of SPIO nanoparticles; both detected through a magnetic probe. Materials & methods: Ten patients with biopsy-proven nonpalpable invasive breast carcinoma or premalignant lesions eligible for SLNs biopsy were enrolled in this study. RESULTS: All tumors were removed with safe surgical margins. The mean nodal detection rate was 3.4 nodes per patient. No interferences in magnetic probe measurements due to the presence of both markers in the same breast were observed. CONCLUSION: Simultaneous use of the magnetic method in localization of impalpable breast tumors and SNs detection makes breast surgery convenient.

Pseudomalignant myositis ossificans in the breast: A case report.

Myositis ossificans (MO) is characterized by abnormal heterotopic ossification formation, typically involving muscles, tendons, ligaments, fascia, and aponeurosis. It can be categorized into nonhereditary and hereditary types, with the latter being a distinct entity with a separate pathophysiology and treatment approach. The pathophysiology of MO formation remains to be fully elucidated. MO is most commonly observed in muscle tissue as a solitary lesion. The disease has been reported to occur in all ages, including the very young and in atypical locations, including hands, feet, ribs, head and neck. The present case report describes an unusual pseudomalignant form of MO in the breast. The authors discuss the clinical and morphological characteristics of the tumor and its treatment options.

Desmoid-type fibromatosis of the breast: A report of 2 cases.

Mammary fibromatosis is a rare and locally aggressive benign tumor of the breast; it originates from fibroblasts and myofibroblasts within the breast parenchyma and does not metastasize. The condition is locally aggressive and has a high rate of recurrence. The etiology of mammary fibromatosis is unknown. Breast imaging examinations are not specific for fibromatosis and often imitate breast cancer. The current study presents 2 cases of women with breast fibromatosis, the first of which exhibited a locally advanced aggressive form of the disease, where breast surgery and en bloc resection of the underlying regions of the thoracic wall were required. In the second case, breast imaging examinations suggested an invasive breast tumor, probably carcinoma, infiltrating the muscles of the chest wall. An ultrasound-guided core needle biopsy revealed a low-grade myofibroblastic proliferation consistent with breast fibromatosis. The patient underwent a right quadrantectomy, with a partial resection of the underlying musculature. The patients remain disease-free at the time of writing. As involvement of the breast in patients with desmoid-like fibromatosis as rare, the present study reports 2 cases with clinical features and histological findings in order to improve and add to the knowledge of this disease.

Dermatofibrosarcoma protuberans of the breast: A case report.

Dermatofibrosarcoma protuberans (DFSP) is a rare malignant tumor of subcutaneous tissue characterized by slow infiltrative growth. The tumor occurs in patients of all ages, with the highest frequency occurring between the second and the fifth decades of age. Genetically, DFSP is characterized by a reciprocal translocation t(17;22)(q22;q13), or more often, as a supernumerary ring chromosome involving chromosomes 17 and 22. Standard treatment of a localized tumor is surgical excision with wide margins. In the present study, a case report of a 43-year-old woman with a growing tumor in the left breast is discussed. The patient underwent breast-conserving surgery. Histological and cytogenetic examinations of the tumor resulted in a diagnosis of DFSP. The clinical and morphological characteristics of the tumor, in addition to the treatment options, were also evaluated.

Cadherin-11 expression is upregulated in invasive human breast cancer.

Loss of expression of cadherin-11 protein is correlated with a loss of epithelial phenotype and a gain in tumor cell proliferation and invasion. It has been hypothesized that cadherin-11 may be a molecular marker for a more aggressive subtype of breast cancer. The present study examined the expression of the mesenchymal gene/protein cadherin-11 in malignant, benign and healthy breast cancer samples. A paraffin-embedded tissue microarray of both malignant and benign/healthy breast tumor was used. Clinicopathological parameters, including age, grading, tumor size, hormone receptors and HER2 receptors status were obtained from patient medical records. Expression of cadherin-11 was analyzed using the monoclonal mouse anti cadherin-11 IgG2B clone. Total RNA was extracted from each breast cancer sample and subjected to semi-quantitative RT-PCR analysis for cadherin-11. Cadherin-11 was detected in 80/82 malignant breast cancer samples and in 33/70 non-malignant tissue samples. Cadherin-11 expression was observed to be predominantly localized to the membrane of tumor cells. When compared to healthy breast tissue biopsies, both cadherin-11 mRNA and protein were demonstrated to be significantly overexpressed in breast carcinoma (P=0.040 and P<0.0001, respectively). Within malignant tumors, however, protein expression was not identified to be associated with other clinicopathological parameters. Our results indicate that cadherin-11 expression is upregulated in malignant human breast cancer.

High-Grade Neuroendocrine Carcinoma of the Breast With Focal Squamous Differentiation.

Primary neuroendocrine carcinoma of the breast is a rare tumor that comprises less than 1% of breast carcinomas, with most patients being in the sixth or seventh decade of their life. In this article, the authors present the case report of a 42-year-old woman with a rapidly growing tumor in her right breast. After clinical staging by physical examination, breast imaging, and thoracoabdominal computed tomography the patient underwent breast-conserving surgery. The histologic results showed a unique type of high-grade, predominantly large-cell neuroendocrine carcinoma with focal abrupt squamous differentiation. The authors also discuss the clinical and morphological characteristics of the tumor as well as treatment options.

Expression of carbonic anhydrase IX in breast is associated with malignant tissues and is related to overexpression of c-erbB2.

CA IX is a tumour-associated carbonic anhydrase with proposed roles in pH modulation and intercellular communication. Its distribution was examined in normal, benign and malignant breast tissues and compared with expression of breast tumour markers including oestrogen receptor, c-erbB2, c-erbB3 and CD44. Tissue specimens were analysed using immunohistochemistry and/or reverse transcriptase-polymerase chain reaction (RT-PCR). CA IX was detected by IHC in 12/26 (46%) malignant tissues, 4/36 (11%) benign lesions, but not in 10 normal breasts. Staining was mostly confined to plasma membranes of abnormal epithelial cells, but in five cases was found in adjacent stroma. Semi-quantitative RT-PCR detected CA9 mRNA in 25/39 (64%) malignant tumours, 11/33 (33%) benign lesions, but in none of three normal breasts. Comparative RT-PCR analysis of malignant tissues revealed a relationship between CA9 positivity and c-erbB2 overexpression (p=0.05). Moreover, CA9-positive specimens displayed a significantly higher median level of c-erbB2 than CA9-negative ones (p=0.02). No significant association was found with the other markers. The results of this study support the possible importance of CA IX for breast carcinogenesis and suggest its potential use as a breast tumour marker.