Detection of Putative Stem-cell Markers in Invasive Ductal Carcinoma of the Breast by Immunohistochemistry: Does It Improve Prognostic/Predictive Assessments?

INTRODUCTION: Experimental evidences from the last 2 decades supports the existence of a special type of neoplastic cell with stem-like features [cancer stem cell (CSC)] and their role in the pathophysiology and therapeutic resistance of breast cancer. However, their clinical value in human breast cancer has not been fully determined. MATERIALS AND METHODS: An immunohistochemistry panel of 10 putative CSC markers (CD34, C-KIT, CD10, SOX-2, OCT 3/4, p63, CD24, CD44, CD133, and ESA/EPCAM) was applied to 74 cases of breast cancer, followed in a Regional Cancer Center of Minas Gerais State, Brazil, from 2004 to 2006. Possible associations between CSC markers and classic variables of clinicopathologic relevance were investigated. RESULTS: The most frequently positive CSC markers were CD44, CD24, CD133, and ESA (the others were present in <15% of the cases). Two CSC profiles were defined: CD24/CD44 (CSC-1) and CD133/ESA (CSC-2). CSC-1 was significantly associated to patients older than 40 years, tumors of <2.0 cm in diameter, early clinical stages (P<0.05), and increased death risk of 4 times (P=0.03; 95% confidence interval, 1.09-14.41). CSC-2 was related to increased relapse risk of 3.75 times (P=0.04; 95% confidence interval, 1.02-13.69). CONCLUSION: The detection of the most frequently positive CSC markers by immunohistochemistry is of clinicopathologic and prognostic relevance.

Tissue-Associated Bacterial Alterations in Rectal Carcinoma Patients Revealed by 16S rRNA Community Profiling.

Sporadic and inflammatory forms of colorectal cancer (CRC) account for more than 80% of cases. Recent publications have shown mechanistic evidence for the involvement of gut bacteria in the development of both CRC-forms. Whereas, colon and rectal cancer have been routinely studied together as CRC, increasing evidence show these to be distinct diseases. Also, the common use of fecal samples to study microbial communities may reflect disease state but possibly not the tumor microenvironment. We performed this study to evaluate differences in bacterial communities found in tissue samples of 18 rectal-cancer subjects when compared to 18 non-cancer controls. Samples were collected during exploratory colonoscopy (non-cancer group) or during surgery for tumor excision (rectal-cancer group). High throughput 16S rRNA amplicon sequencing of the V4-V5 region was conducted on the Ion PGM platform, reads were filtered using Qiime and clustered using UPARSE. We observed significant increases in species richness and diversity in rectal cancer samples, evidenced by the total number of OTUs and the Shannon and Simpson indexes. Enterotyping analysis divided our cohort into two groups, with the majority of rectal cancer samples clustering into one enterotype, characterized by a greater abundance of Bacteroides and Dorea. At the phylum level, rectal-cancer samples had increased abundance of candidate phylum OD1 (also known as Parcubacteria) whilst non-cancer samples had increased abundance of Planctomycetes. At the genera level, rectal-cancer samples had higher abundances of Bacteroides, Phascolarctobacterium, Parabacteroides, Desulfovibrio, and Odoribacter whereas non-cancer samples had higher abundances of Pseudomonas, Escherichia, Acinetobacter, Lactobacillus, and Bacillus. Two Bacteroides fragilis OTUs were more abundant among rectal-cancer patients seen through 16S rRNA amplicon sequencing, whose presence was confirmed by immunohistochemistry and enrichment verified by digital droplet PCR. Our findings point to increased bacterial richness and diversity in rectal cancer, along with several differences in microbial community composition. Our work is the first to present evidence for a possible role of bacteria such as B. fragilis and the phylum Parcubacteria in rectal cancer, emphasizing the need to study tissue-associated bacteria and specific regions of the gastrointestinal tract in order to better understand the possible links between the microbiota and rectal cancer.

An Integrative Approach Uncovers Biomarkers that Associate with Clinically Relevant Disease Outcomes in Vulvar Carcinoma.

UNLABELLED: Vulvar squamous cell carcinoma (VSCC) is a rare disease that has a high mortality rate (∼40%). However, little is known about its molecular signature. Therefore, an integrated genomics approach, based on comparative genome hybridization (aCGH) and genome-wide expression (GWE) array, was performed to identify driver genes in VSCC. To achieve that, DNA and RNA were extracted from frozen VSCC clinical specimens and examined by aCGH and GWE array, respectively. On the basis of the integration of data using the CONEXIC algorithm, PLXDC2 and GNB3 were validated by RT-qPCR. The expression of these genes was then analyzed by IHC in a large set of formalin-fixed paraffin-embedded specimens. These analyses identified 47 putative drivers, 46 of which were characterized by copy number gains that were concomitant with overexpression and one with a copy number loss and downregulation. Two of these genes, PLXDC2 and GNB3, were selected for further validation: PLXDC2 was downregulated and GNB3 was overexpressed compared with non-neoplastic tissue. By IHC, both proteins were ubiquitously expressed throughout vulvar tissue. High expression of GNB3 and low PLXDC2 immunostaining in the same sample was significantly associated with less lymph node metastasis and greater disease-free survival. On the basis of a robust methodology never used before for VSCC evaluation, two novel prognostic markers in vulvar cancer are identified: one with favorable prognosis (GNB3) and the other with unfavorable prognosis (PLXDC2). IMPLICATIONS: This genomics study reveals markers that associate with prognosis and may provide guidance for better treatment in vulvar cancer. Mol Cancer Res; 14(8); 720-9. ©2016 AACR.

Breast Carcinoma-associated Fibroblasts Share Similar Biomarker Profiles in Matched Lymph Node Metastasis.

This study sought to understand the role of breast carcinoma-associated fibroblasts in the progression of cancer cells into lymph nodes. We compared fibroblasts of primary tumors and matched the involved lymph nodes to select fibroblast activation markers, namely α-smooth muscle actin (α-SMA), S100A4, and vimentin, as well as to determine the frequency of transforming growth factor ß1, a pleiotropic cytokine that induces the differentiation of fibroblasts to myofibroblasts, and its downstream effectors: CXCR4 and p-AKT. We disposed samples of 80 primary invasive ductal carcinomas and matched the involved lymph nodes from 43 cases into 3 tissue microarrays, and analyzed stromal and tumor epithelial cells separately by immunohistochemistry. Control uninvolved lymph nodes were analyzed by whole-tissue sections. Cancer-associated fibroblast in lymph nodes with macrometastasis expressed similar profiles of vimentin, α-SMA, and S100A4 as those found in primary tumors. Cancer-associated fibroblast were uniformly estrogen receptor, progesterone receptor, HER-2, Ki-67, and p53 negative, but expressions of transforming growth factor ß1 (TGFß1), CXCR4, and p-AKT staining (62.3%, 52.4%, 65%, respectively) were equivalent between primary and lymph node metastasis (LNM) fibroblasts. A significant coexpression of TGFß1 with p-AKT and CXCR4 in LNMs suggested the involvement of these proteins with TGFß1 signaling. These biomarkers, including α-SMA and S100A4, were negative in fibroblasts of cancer-free lymph nodes, with the exception of vimentin. Our finding that expressions of biological markers were similar in fibroblasts of the primary tumors and in matched LNMs, but were absent in cancer-free lymph nodes, supports the assumption that the lymph node stroma mimics the microenvironment observed in primary tumors.

Design of a miRNA sponge for the miR-17 miRNA family as a therapeutic strategy against vulvar carcinoma.

Dysregulation of microRNAs has been studied thoroughly, and has been observed in a variety of tumors including vulvar carcinomas, a rare type of gynecological tumor with increasing incidence. However, very few therapeutic alternatives have reached the clinical setting, and there is an urgent unmet need to develop novel strategies for patients with this tumor type. Thus, a microRNA (miRNA) sponge for the miR-17 miRNA family was designed, synthesized and validated in vitro in order to explore a new therapeutic strategy based on inhibiting this oncogenic miRNA family in vulvar cancer. Members of the miR-17 family were evaluated for expression in a vulvar tumor cell line (SW954) and 20 HPV negative formalin-fixed paraffin-embedded (FFPE) samples by quantitative real-time PCR (qRT-PCR). Six in tandem, bulged sequences that were complementary to these miRNAs were designed, synthesized, cloned, and transfected into SW954 cells. A luciferase reporter assay with a psiCheck2 vector was used to test the specificity of the sponge sequences for miR-17 family miRNA binding. Taqman qRT-PCR was used to test how the sponges affected miRNA expression. In FFPE samples, higher expression of miR-20a and miR-106a correlated with deeper tumor invasion (P = 0.0187 and P = 0.0404, respectively). The luciferase reporter assay validated the specificity of the sponge for miR-17 family members. Using qRT-PCR, we confirmed this specificity with decreased expression in 5 (out of six) miRNAs of the miR-17 family in SW954 cells. Although our results are preliminary, these results demonstrate that these miRNA sponges are potent inhibitors of the miR-17 family of miRNAs in SW954. Therefore, this miRNA-specific sponge may be developed into a novel therapeutic treatment for patients with vulvar cancer.

Prostate-specific RNA aptamer: promising nucleic acid antibody-like cancer detection.

We described the selection of a novel nucleic acid antibody-like prostate cancer (PCa) that specifically binds to the single-stranded DNA molecule from a 277-nt fragment that may have been partially paired and bound to the PCA3 RNA conformational structure. PCA3-277 aptamer ligands were obtained, and the best binding molecule, named CG3, was synthesized for validation. Aiming to prove its diagnostic utility, we used an apta-qPCR assay with CG3-aptamer conjugated to magnetic beads to capture PCA3 transcripts, which were amplified 97-fold and 7-fold higher than conventional qPCR in blood and tissue, respectively. Histopathologic analysis of 161 prostate biopsies arranged in a TMA and marked with biotin-labeled CG3-aptamer showed moderate staining in both cytoplasm and nucleus of PCa samples; in contrast, benign prostatic hyperplasia (BPH) samples presented strong nuclear staining (78% of the cases). No staining was observed in stromal cells. In addition, using an apta-qPCR, we demonstrated that CG3-aptamer specifically recognizes the conformational PCA3-277 molecule and at least three other transcript variants, indicating that long non-coding RNA (lncRNA) is processed after transcription. We suggest that CG3-aptamer may be a useful PCa diagnostic tool. In addition, this molecule may be used in drug design and drug delivery for PCa therapy.

ROCK1 as a novel prognostic marker in vulvar cancer.

BACKGROUND: Vulvar carcinoma is an infrequent tumour, accounting for fewer than 3% of all malignant tumours that affect women, but its incidence is rising in the past few decades. In young women, the manifestation of the vulvar carcinoma is often linked to risk factors such as smoking and HPV infection, but most cases develop in women aged over 50 years through poorly understood genetic mechanisms. Rho-associated coiled-coil-containing protein kinase 1 (ROCK1) has been implicated in many cellular processes, but its function in vulvar cancer has never been examined. In this study, we aimed to determine the prognostic value of ROCK1 gene and protein analysis in vulvar squamous cell carcinoma (VSCC). METHODS: ROCK1 expression levels were measured in 16 vulvar tumour samples and adjacent normal tissue by qRT-PCR. Further, 96 VSCC samples were examined by immunohistochemistry (IHC) to confirm the involvement of ROCK1 in the disease. The molecular and pathological results were correlated with the clinical data of the patients. Sixteen fresh VSCC samples were analyzed by array-based comparative genomic hybridization (aCGH). RESULTS: In each pair of samples, ROCK1 levels were higher by qRT-PCR in normal tissue compared with the tumour samples (p = 0.016). By IHC, 100% of invasive front areas of the tumour and 95.8% of central tumour areas were positive for ROCK1. Greater expression of ROCK1 was associated with the absence of lymph node metastasis (p = 0.022) and a lower depth of invasion (p = 0.002). In addition, higher ROCK1 levels correlated with greater recurrence-free survival (p = 0.001). Loss of ROCK1 was independently linked to worse cancer-specific survival (p = 0.0054) by multivariate analysis. This finding was validated by IHC, which demonstrated enhanced protein expression in normal versus tumour tissue (p < 0.001). By aCGH, 42.9% of samples showed a gain in copy number of the ROCK1 gene. CONCLUSIONS: ROCK1 is lower expressed in tumour tissue when compared with adjacent normal vulvar epithelia. In an independent sample set of VSCCs, lower expression levels of ROCK1 correlated with worse survival rates and a poor prognosis. These findings provide important information for the clinical management of vulvar cancer.

PGC-1α mediates mitochondrial biogenesis and oxidative phosphorylation in cancer cells to promote metastasis.

Cancer cells can divert metabolites into anabolic pathways to support their rapid proliferation and to accumulate the cellular building blocks required for tumour growth. However, the specific bioenergetic profile of invasive and metastatic cancer cells is unknown. Here we report that migratory/invasive cancer cells specifically favour mitochondrial respiration and increased ATP production. Invasive cancer cells use the transcription coactivator peroxisome proliferator-activated receptor gamma, coactivator 1 alpha (PPARGC1A, also known as PGC-1α) to enhance oxidative phosphorylation, mitochondrial biogenesis and the oxygen consumption rate. Clinical analysis of human invasive breast cancers revealed a strong correlation between PGC-1α expression in invasive cancer cells and the formation of distant metastases. Silencing of PGC-1α in cancer cells suspended their invasive potential and attenuated metastasis without affecting proliferation, primary tumour growth or the epithelial-to-mesenchymal program. Inherent genetics of cancer cells can determine the transcriptome framework associated with invasion and metastasis, and mitochondrial biogenesis and respiration induced by PGC-1α are also essential for functional motility of cancer cells and metastasis.

A unified nomenclature and amino acid numbering for human PTEN.

The tumor suppressor PTEN is a major brake for cell transformation, mainly due to its phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P3] phosphatase activity that directly counteracts the oncogenicity of phosphoinositide 3-kinase (PI3K). PTEN mutations are frequent in tumors and in the germ line of patients with tumor predisposition or with neurological or cognitive disorders, which makes the PTEN gene and protein a major focus of interest in current biomedical research. After almost two decades of intense investigation on the 403-residue-long PTEN protein, a previously uncharacterized form of PTEN has been discovered that contains 173 amino-terminal extra amino acids, as a result of an alternate translation initiation site. To facilitate research in the field and to avoid ambiguities in the naming and identification of PTEN amino acids from publications and databases, we propose here a unifying nomenclature and amino acid numbering for this longer form of PTEN.

Immunoexpression of S100A4 in canine skin melanomas and correlation with histopathological parameters.

BACKGROUND: Melanoma is one of the most common skin neoplasms in humans and dogs. The tumor microenvironment in melanoma comprises cancer cells and stromal cells that interact to accelerate tumor progression. Several prognostic markers for melanomas have been studied in many human tumors, including fibroblast-specific protein 1 (S100A4). S100A4 is a member of the S100 family of calcium-binding proteins in stromal cells. HYPOTHESIS/OBJECTIVES: The objective of this study was to describe the immunohistochemical patterns of S100A4 in stroma and neoplastic cells of canine skin melanomas and correlate them with some histological parameters. ANIMALS AND METHODS: Forty-eight samples (38 pigmented and 10 non-pigmented melanomas) were first selected and their nature confirmed using S100, Melan A and vimentin. All cases were examined by immunohistochemistry using S100A4 to correlate expression, histotype, and level of invasion. RESULTS: All the tumors, including 10 non-pigmented, were positive for S100, Melan A, vimentin and negative for cytokeratin AE1/AE3 (consistent with melanomas). The 48 melanomas were classified as epithelioid (n = 21), spindle (n = 14), and mixed (n = 13). S100A4 was preferentially expressed in epithelioid and spindle cell types compared with mixed melanomas and S100A4 expression was not associated with level of invasion (Clark's levels IV to V). CONCLUSION: S100A4 expression in melanoma samples varied among histotypes but not between levels of invasion.

Dynamic dialog between cytokeratin 18 and annexin A1 in breast cancer: a transcriptional disequilibrium.

Cytokeratins (CKs) constitute the cytoskeletal network and are regulated by post-translational modifications, acting not only as a mechanical support, but also in cell signaling and regulatory processes. Signaling is mediated by CK-associated proteins, such as Annexin A1 (ANXA1), a ligand of the CK18/CK8 complex. ANXA1 has a pivotal role in cellular and immunological responses, and together with CK18 have been implicated in several processes related to malignant transformation in breast cancer (BC). Our aim was to demonstrate how their interaction might be linked to BC development. We investigated transcript levels, protein expression and distribution for both targets in breast tissues of 92 patients (42 BCs and 50 benign diseases) using qPCR and immunohistochemistry, respectively. ANXA1 and CK18 mRNAs were inversely correlated, and their ratio in each TNM stage significantly differentiated BC from benign diseases (OR=5.62). These differences did not mirror tissue protein levels, but a significant dichotomous protein distribution in tumor tissues was observed, differing from the expected co-localization observed during cell homeostasis. The disequilibrium of transcriptional levels between ANXA1/CK18 and alterations in their tissue distribution are present either in initial events or tumor progression, which suggest a critical event in BC. The broken dialog between ANXA1 and CK18 in normal breast tissues may play a critical role in BC development, and together may be used as combined targets for BC diagnostics.

Repair of oxidative DNA damage, cell-cycle regulation and neuronal death may influence the clinical manifestation of Alzheimer's disease.

Alzheimer's disease (AD) is characterized by progressive cognitive decline associated with a featured neuropathology (neuritic plaques and neurofibrillary tangles). Several studies have implicated oxidative damage to DNA, DNA repair, and altered cell-cycle regulation in addition to cell death in AD post-mitotic neurons. However, there is a lack of studies that systematically assess those biological processes in patients with AD neuropathology but with no evidence of cognitive impairment. We evaluated markers of oxidative DNA damage (8-OHdG, H2AX), DNA repair (p53, BRCA1, PTEN), and cell-cycle (Cdk1, Cdk4, Cdk5, Cyclin B1, Cyclin D1, p27Kip1, phospho-Rb and E2F1) through immunohistochemistry and cell death through TUNEL in autopsy hippocampal tissue samples arrayed in a tissue microarray (TMA) composed of three groups: I) "clinical-pathological AD" (CP-AD)--subjects with neuropathological AD (Braak ≥ IV and CERAD = B or C) and clinical dementia (CDR ≥ 2, IQCODE>3.8); II) "pathological AD" (P-AD)--subjects with neuropathological AD (Braak ≥ IV and CERAD = B or C) and without cognitive impairment (CDR 0, IQCODE<3.2); and III) "normal aging" (N)--subjects without neuropathological AD (Braak ≤ II and CERAD 0 or A) and with normal cognitive function (CDR 0, IQCODE<3.2). Our results show that high levels of oxidative DNA damage are present in all groups. However, significant reductions in DNA repair and cell-cycle inhibition markers and increases in cell-cycle progression and cell death markers in subjects with CP-AD were detected when compared to both P-AD and N groups, whereas there were no significant differences in the studied markers between P-AD individuals and N subjects. This study indicates that, even in the setting of pathological AD, healthy cognition may be associated with a preserved repair to DNA damage, cell-cycle regulation, and cell death in post-mitotic neurons.

Vulvar cancer surgery.

PURPOSE OF REVIEW: Surgical treatment of vulvar cancer has been shifted from ultraradical procedures associated with huge morbidity to less extensive surgery with better psychosexual result and less morbidity, without compromising survival. The authors review and discuss the recent literature regarding the surgical management of vulvar squamous cell carcinoma. RECENT FINDINGS: Surgery remains the cornerstone in the treatment of vulvar cancer. Radical vulvectomy with inguinofemoral lymphadenectomy has been replaced by radical local excision with sentinel node procedure for early disease. However, the role and distance of pathological margins are still on debate. Recent results from a large prospective trial corroborate the safety of sentinel node biopsy for early disease, even after primary tumor resection. An experienced team should perform sentinel node procedure using combined technique (blue dye and lymphoscintigraphy) and ultrastaging pathology. Moreover, midline tumors still need lymph node biopsy from both groins. SUMMARY: Primary vulvar cancer may be safely treated with radical/wide local resection. In case of other suspicious lesion or multifocal disease, radical vulvectomy is performed. Patients with unifocal disease, tumor size less than 4 cm, and clinically negative groins are candidates to sentinel node procedure. In the case of clinically positive node or sentinel node metastasis, a systematic inguinofemoral lymphadenectomy should be performed.

Clinical significance of the interaction between non-coding RNAs and the epigenetics machinery: challenges and opportunities in oncology.

Non-coding RNAs and epigenetics are remarkable mechanisms of cellular control. In this review we underline the processes by which non-coding RNAs (ncRNAs), shown to be involved in various diseases, are capable of modifying and being modified by the epigenetic machinery, emphasizing the clinical importance of this network in cancer. Many ncRNAs have been described that play important roles in the establishment and maintenance of the epigenome. However, only a few studies deeply take into account the role of ncRNAs from a clinicopathological standpoint. The wide range of interactions between the non-coding RNome and the epigenome, and the roles of these networks in the pathogenesis, prognosis and early diagnosis of many diseases, present new challenges and opportunities for future studies regarding therapeutic strategies in oncology.

A novel highly reactive Fab antibody for breast cancer tissue diagnostics and staging also discriminates a subset of good prognostic triple-negative breast cancers.

The discovery of novel markers for breast cancer (BC) has been recently relied on antibody combinatorial libraries and selection through phage display. We constructed a recombinant Fab library, and after selections against BC tissues, the FabC4 clone was thoroughly investigated by immunohistochemistry in 232 patients with long-term follow-up. The FabC4 ligand was determined by mass spectrometry. The FabC4 expression was associated with younger age, lack of progesterone receptor, higher histological grades and non-luminal subtypes, and it also identified a subset of good prognostic triple-negative BCs, possibly targeting a conformational epitope of Cytokeratin-10 (CK10). This new CK10-epitope specific antibody may open new possibilities in diagnostic and therapeutic strategies.

Best practice for PTEN gene and protein assessment in anatomic pathology.

There is a lack of standardization of a best practice protocol for Phosphatase and Tensin Homolog (PTEN) assessment by immunohistochemistry in anatomic pathology routine practice. We performed immunohistochemistry for 19 antibodies against PTEN, eleven of which were excluded during the standardization step. Immunohistochemistry of the remaining eight antibodies was performed on a Tissue Microarray containing 55 prostate and 40 renal carcinoma samples. Fluorescent in situ hybridization (FISH) was used as reference standard for immunohistochemistry specificity evaluation. Concerning nuclear staining, polyclonal (Cat#22034-1-AP); 6H2.1 mMAb (Cat#ABM-2052), Y184 RabMAb (Cat#NB110-57441) and 217702 mMAb antibodies presented the highest agreement with fluorescent in situ hybridization (p<0.001 for all) and with regard to cytoplasmic staining, Y184 RabMAb (Cat#NB110-57441); polyclonal (Cat#22034-1-AP) and 217702 mMAb presented the highest agreement (p<0.001 for all). Our results indicate that several commercially available antibodies do not show reliability of sensitivity and specificity for PTEN evaluation and we propose 6H2.1 mMAb (Cat#ABM-2052) as the antibody of choice for laboratory standardization and best practice in clinical routine, which demonstrated excellent sensitivity for both nuclear and cytoplasmic staining, specificity for PTEN by Western blot and good correlation with PTEN status by FISH with regard to nuclear staining.

Polybromo-1 (PBRM1), a SWI/SNF complex subunit is a prognostic marker in clear cell renal cell carcinoma.

OBJECTIVE: To analyse the immunohistochemical and mRNA expression of SWI/SNF (SWItch/Sucrose NonFermentable) complex subunit polybromo-1 (PBRM1) in clear cell renal cell carcinoma (ccRCC) and its impact on clinical outcomes. PATIENTS AND METHODS: In all, 213 consecutive patients treated surgically for renal cell carcinoma (RCC) between 1992 and 2009 were selected. A single pathologist reviewed all cases to effect a uniform reclassification and determined the most representative tumour areas for construction of a tissue microarray. In addition, mRNA expression of PBRM1 was analysed by reverse transcriptase-polymerase chain reaction. RESULTS: Of the 112-immunostained ccRCC specimens, 34 (30.4%) were PBRM1-negative, and 78 (69.6%) were PBRM1-positive. The protein expression of PBRM1 was associated with tumour stage (P < 0.001), clinical stage (P < 0.001), pN stage (P = 0.035) and tumour size (P = 0.002). PBRM1 mRNA expression was associated with clinical stage (P = 0.023), perinephric fat invasion (P = 0.008) and lymphovascular invasion (P = 0.042). PBRM1 significantly influenced tumour recurrence and tumour-related death. Disease-specific survival rates for patients whose specimens showed positive- and negative-PBRM1 expression were 89.7% and 70.6%, respectively (P = 0.017). Recurrence-free survival rates in patients with positive- and negative-expression of PBRM1 were 87.3% and 66.7%, respectively (P = 0.048). CONCLUSIONS: PBRM1-negative expression is a markedly poor prognosis event in ccRCC. We encourage PBRM1 study by other groups in order to validate our findings and confirm its possible role as a useful marker in the management of patients with ccRCC.

Clinical, cytologic, and histologic features of a mammary micropapillary carcinoma in a dog.

Mammary invasive micropapillary carcinoma is a rare variant of mammary carcinoma that was recently recognized in dogs. The cytologic features and biologic behavior of such neoplasms in dogs have not yet been widely discussed in the veterinary literature. We report the clinical, cytologic, and histologic features of a canine micropapillary carcinoma in a 13-year-old female mongrel dog. The mammary region presented with extreme local pain, severe edema and erythema, and multifocal epidermal ulceration, which is typical for an inflammatory mammary carcinoma. Fine-needle aspirates were highly cellular and consisted of individual cells and papillary cell clusters with characteristics of malignant epithelial cells. Histologic examination revealed neoplastic cells arranged in small papillae without fibrovascular cores, sometimes inside clear lymphatic spaces, indicating lymphovascular invasion. Regional lymph node evaluation revealed metastatic cells. Due to deteriorating clinical condition the dog was euthanatized 5 months after mastectomy. At necropsy, metastatic neoplastic mammary cells were found in popliteal and mediastinal lymph nodes, the right femoral biceps muscle, liver, heart, lungs, and urinary bladder.

Usefulness and limitations of E-cadherin and ß-catenin in the classification of breast carcinomas in situ with mixed pattern.

BACKGROUND: The distinction between lobular neoplasia of the breast and ductal carcinoma in situ has important therapeutic implications. In some cases, it is very difficult to determine whether the morphology of the lesion is ductal or lobular. The aim of this study was to evaluate the value of E-cadherin and ß-catenin expression through the immunophenotypical characterization of carcinoma in situ with mixed pattern (CISM). METHODS: A total of 25 cases of CISM were analyzed considering cytology/mixed architecture (ductal and lobular), nuclear pleomorphism, loss of cell cohesion, and presence of comedonecrosis. The immunophenotype pattern was considered E-cadherin positive and ß-catenin positive, or negative. RESULTS: Nineteen (76%) cases presented a mixed cytology and / or architectural pattern, two (8%) presented nuclear pleomorphism, two (8%) presented mixed cytology and nuclear pleomorphism, and two (8%) presented comedonecrosis and nuclear pleomorphism. A complete positivity for E-cadherin and ß-catenin was observed in 11 cases (44%). In one case, the lesion was negative for both markers and showed nuclear pleomorphis. Thirteen lesions showed negative staining in areas of lobular cytology and positive staining in cells presenting the ductal pattern. CONCLUSIONS: The expression of E-cadherin and ß-catenin, combined with cytological and architectural analysis, may highlight different immunophenotypes and improve classification of CISM. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/ 1693384202970681

Relação da expressão de fatores de crescimento celular (IGF-1) e (SCF) com fatores prognósticos e o alvo da rapamicina em mamíferos (m-TOR) em mastocitomas cutâneos caninos / IGF-1 and SCF protein expression in cutaneous mast cell tumors in dogs and relation to prognostic factors and mammalian target of rapamycin (m-TOR)

O mastocitoma cutâneo (MTC) é a neoplasia maligna mais comum na pele dos cães e seu comportamento biológico é muito variável. Dentre os fatores prognósticos estudados nos MTCs, a classificação histopatológica, o índice proliferativo e o padrão de expressão doc-KIT são os que apresentam uma associação mais relevante com o provável prognóstico deste tumor. O objetivo deste trabalho foi avaliar a expressão proteica de fator de crescimento semelhante à insulina tipo 1 (IGF-1), fator de célula tronco (SCF) e sua relação com o receptor tirosina quinase (c-KIT), alvo da rapamicina em mamíferos (m-TOR), grau histológico, índice proliferativo pelo KI-67e o número de figuras de mitose (IM) com dados clínicos de cães com MTCs . Foram utilizadas 133 amostras de MTCs, provenientes de 133 cães, dispostas em lâminas de microarranjo de tecidos (TMA). A técnica de imuno-histoquímica foi utilizada para a avaliação destas proteínas. Observou-se associação entre SCF e, a graduação histopatológica proposta em 2011, índice mitótico, proliferação celular (KI-67), escore de IGF-1, local da lesão, idade dos animais e padrão imuno-histoquímico do receptor c-KIT. A relação de dependência também foi observada entre IGF-1 e o porte dos animais, IM, m-TOR e c-KIT. A expressão de SCF teve relacção com a agressividade dos MTCs caninos, uma vez que foi mais freqüente em MTCs com c-KIT citoplasmático. A relação entre a expressão de IGF-1, SCF, c-KIT e m-TOR pode estar associada à integralização de suas vias de ação. A expressão de IGF-1 está associada à MTCs em cães de porte grande.

Cutaneous mast cell tumor (MCT) is one of the most common neoplasms in the skin of dogs and express variable biological behavior. Among the MTC aspects studied, histological classification, proliferative index and protein expression of c-KIT show the most defined connection with the tumor prognostic. The aim of this study was to evaluate the protein expression of insulin-like growth factor type 1 (IGF-1), steam cell factor (SCF) and theit relationship with tyrosine kinase receptor (c-KIT), mammalian target of rapamycin (m-TOR), histological classification (KI-67), proliferative and mitotic index and epidemiological data in MTCs. In this study 133 MTC samples from 133 animals were used, arranged in tissue microarray (TMA) slides. The TMA was used for evaluation the proteins. An association was observed between SCF and histological grade proposed in 2011, mitotic index, cell proliferation, IGF-1, lesion site, age of the animals, and immunohistochemical pattern c-KIT receptor. The dependence relationship was also observed between IGF-1 and animal size, mitotic index, m-TOR and c-KIT. The SCF protein expression was related to canine MTCs aggressiveness, since it is more frequent in MCTs with c-KIT cytoplasmic. The relationship between the expression of IGF-1, SCF, c-KIT e m-TOR can be associated with the integration of its actions ways. The IGF-1 expression is associated with large dog breeds MTCs.