We report two new cases of cystic fi broepithelioma of Pinkus together with immunohistochemical features and analyze the presence of cystic changes in a series of 16 classical fi broepitheliomas of Pinkus. Our fi ndings show that the formation of cystic spaces is most probably caused by ischemic degeneration of stromal fenestrations, rather than by central tumor cell necrosis. Th is fi nding is supported by lack of CD34 positive blood vessels in edematous and hyalinized stromal fenestrations undergoing transformation into cystic spaces, as opposed to the uninvolved stromal fenestrations. Th erefore, it is probably more accurate to refer to this process as pseudocystic stromal degeneration rather than true cyst formation. Also, two out of 16 classical Pinkus fi broepitheliomas exhibited focal pseudocystic changes in 50 and 10 of the tumor, respectively, demonstrating that this degenerative process can be found, rarely and focally, in classical cases as well.

Metastases to gastrointestinal tract are uncommon. In particular, metastases to the ampulla of Vater are very rare and may represent a significant diagnostic challenge. Metastases from the uterine cervix to the ampulla of Vater are exceedingly rare and only one case has been described in the available literature. We describe here a second case of metastatic squamous cell carcinoma of the cervix to the ampulla of Vater in a 45-year-old woman. Poorly differentiated squamous cell carcinoma presented as an isolated metastasis to the ampulla of Vater, two years after the initial diagnosis. While the squamous cell carcinoma could occur as primary ampullary carcinoma, albeit very rare, it is necessary to exclude the possibility of metastatic cancer.

Apocrine carcinoma of the breast is a rare, special type of breast carcinoma showing distinct morphologic, immunohistochemical and molecular genetic features. Apocrine epithelium has a characteristic steroid receptor profile that is estrogen receptor and progesterone receptor negative and androgen receptor positive. This combination of morphologic and immunohistochemical characteristics is essential for the proper recognition of the apocrine carcinomas. Strictly defined, apocrine carcinomas express either Her-2/neu or EGFR, which along with androgen receptor positivity make patients with the apocrine carcinoma eligible for targeted therapies.

Apocrine differentiation occurs in a variety of breast lesions, both benign and malignant. The molecular classification of breast cancer defined an apocrine molecular subtype of breast cancer. In general, apocrine carcinomas do not present clinical behavior distinct from the observed in ductal carcinomas of no special type. However, these carcinomas are frequently triple negative (ER-/PgR-/HER2-) with no targeted therapy and poor prognosis. Still they are unique in the aspect of the expression of a different nuclear receptor - androgen receptor (AR). In this study we thought to perform a comprehensive genetic/molecular characterization of this type of lesions. For that, a series of 44 apocrine lesions were studied. These comprised 9 apocrine metaplasia, 8 apocrine adenosis, 13 apocrine in situ carcinoma and 18 apocrine invasive carcinoma. Importantly four samples contained different lesions in different stages in close proximity. Immunohistochemistry was performed to evaluate the ER, PgR, HER2 and AR status of these lesions. Genomic DNA was microdissected from FFPE slides and used both to identify chromosomal copy number aberrations using aCGH and to monitor 739 mutations from 46 oncogenes and tumor suppressor genes using the Ion PGM system. The aCGH study allowed the identification of recurrent chromosomal aberrations. These were rare in benign apocrine lesions, mostly comprised of small gains and deletions, and dramatically increased in malignant lesions up to complete loss and gain of full chromosome arms. Also the analysis of the different lesions within the same sample indicates a putative existence of an apocrine tumor progression. Moreover the profiling of mutational status of the different tumor suppressor/oncogene allowed the identification of mutations that may provide new avenues for the development of new targetable therapeutically options and also corroborated the putative apocrine tumor progression. The study of apocrine lesions in different differentiation stages allowed a deeper insight on the molecular complexity of these lesions. Although further studies are required to independently validate this work, this study identifies recurrent genomic alteration that may open targeted treatment opportunities that may be used independently or in combination with anti-androgen drugs. Citation Format: Jose L. Costa, Ana Justino, Madalena Gomes, Cesar Augusto Alvarenga, Rene Gerhard, Semir Vranic, Zoran Gatalica, Jose Carlos Machado, Fernando Schmitt. Comprehensive genetic characterization of apocrine lesions of the breast. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2013. doi:10.1158/1538-7445.AM2013-2013

p53 is one of the most frequently mutated genes in human tumors including head and neck tumors like oral squamous cell carcinoma. It might be responsible for more than 50% of all relapses in patients with surgically treated oral carcinoma and clean margins. The aim of the present study was to explore p53 protein expression in peritumoral tissue and correlate it with relapse of the disease. The study included 25 patients (17 males and 8 females) with oral squamous cell carcinoma in the period August 2006 till August 2008. For immunohistochemical assay, a monoclonal antibody against p53 protein was applied (clone DO-7, DAKO Glostrup, Denmark). Peritumoral expression of p53 was as follows: 10 out of 25 cases (40%) were negative, 2 cases (8%) showed weak, 5 cases (20%) moderate and 8 cases (32%) strong p53 positivity. No significant correlation between peritumoral expression of p53 protein and patient's relapse was found. In contrast, we found a trend toward association between intratumoral p53 expression and patient's relapse (p = 0.07). There was also trend toward higher peritumoral p53 expression in females comparing with p53 expression in males (52.9% of males did not have p53 expression while 87.5% females had mild, moderate or high p53 expression, p = 0.088). Peritumoral expression of p53 protein is frequently seen in oral squamous cell carcinoma and merits further research.

The study was undertaken to investigate EGFR and HER-2/neu expression in a cohort of apocrine carcinomas of the breast with emphasis on the classification of the breast carcinomas with apocrine morphology. In total, 55 breast carcinomas morphologically diagnosed as apocrine were evaluated for steroid receptor expression profile characteristic of normal apocrine epithelium (ER-/PR-/AR+), and for the expression of EGFR and Her-2/neu proteins, and the copy number ratios of the genes EGFR/CEP7 and HER-2/CEP17. Another cohort composed of 72 invasive ductal carcinomas of no-special-type was used to further determine the impact of CEP17 polysomy on the interpretation of HER-2/neu testing. Our study confirms that apocrine carcinomas of the breast are molecularly diverse group of carcinomas. Strictly defined, pure apocrine carcinomas (ER-, PR-, AR+) (38 cases, 69%) are either HER-2 overexpressing breast carcinomas (52%) or triple-negative breast carcinomas (48%). Apocrine-like carcinomas (ER+/-, PR+/-, AR+/-) (17 cases, 31%) belong predominantly to the luminal phenotype (76%). Pure apocrine carcinomas show consistent over-expression of either EGFR or Her-2/neu. EGFR gene amplification was observed in two pure apocrine carcinomas and one apocrine-like carcinoma. CEP7 polysomy (defined as three or more CEP7 signals) was seen in 61% pure apocrine carcinomas and 27% of apocrine-like carcinomas and showed a weak positive correlation with EGFR protein expression. HER-2/neu gene amplification is the primary mechanism of Her-2/neu activation and is found in 52% of all apocrine carcinomas. CEP17 polysomy (defined as three or more CEP17 signals) was observed in 10 pure apocrine carcinomas (32%) and 8 apocrine-like carcinomas (50%). CEP17 polysomy may be seen without HER-2/neu gene amplification. Further exploration on a cohort of invasive ductal carcinomas of no-special-type confirmed that increased CEP17 signals may lead to discordant interpretation of HER-2/neu gene amplification in a significant proportion of the cases, depending on which criterion (ratio versus absolute number) is used for interpretation. However, increased gene dosage (>6 HER-2/neu genes or HER-2/CEP17 ratio>2.2), regardless of the evaluation method, is positively correlated with Her-2/neu protein expression.

Cellular hypoxia is a hallmark of cancer. Hypoxia-inducible factor-1&agr; (HIF-1&agr;) and von Hippel-Lindau protein (pVHL) are the key mediators of cellular response to hypoxia. Little is known about their role in germ cell tumors of the testis. We therefore examined their status in a cohort of germ cell tumors of the testis. Thirty-six primary germ cell tumors of the testis (11 seminomas, 24 mixed germ cell tumors, and 1 case of pure intratubular germ cell neoplasia) were included in the study. HIF-1&agr; and pVHL expression were studied using immunohistochemical (IHC) methods in the tumor and adjacent benign tissue. Selected cases with a low pVHL expression were further tested for genetic alterations using polymerase chain reaction. HIF-1&agr; protein expression was not detectable in adjacent atrophic seminiferous tubules. In contrast, HIF-1&agr; was expressed in one third of the malignancies, but in a low percentage of cells (mean, 3%; range, 0% to 20%). No difference in HIF-1&agr; expression was observed between seminomas and nonseminomas (P=0.71). pVHL was expressed in atrophic tubular epithelium and in the Leydig cells, whereas a substantial loss of pVHL expression was observed in germ cell tumors regardless of the histologic type (mean, 45.6%; range, 0% to 100%). No genetic alterations of the VHL gene were observed in the cases with low pVHL expression. No significant correlation between HIF-1&agr; and pVHL expression was observed (P=0.16). Germ cell tumors of the testis, regardless of the histologic type, are characterized by consistently low HIF-1&agr; protein overexpression and a partial loss of pVHL without underlying VHL gene alterations. Further studies are necessary to clarify the functional importance of such alterations in testicular germ cell tumors.

Apocrine carcinoma of the breast has recently been refined through gene expression profiling. Due to various pathological studies, we compared the results with the MDA-MB-453 breast cancer cell line, a proposed model for apocrine breast carcinoma. The MDA-MB-453 cell line is androgen receptor-positive and `triple-negative' in respect to estrogen receptor-α, progesterone receptor and the Her-2/neu protein expression. Cytogenetic analysis of the cell line revealed a hypertriploid clone characterized by extensive numerical and structural abnormalities including loss of the 9p.21 locus (P16-INK4a gene), also evidenced by the lack of p16(INK4A) protein expression in Western blot analysis and immunocytochemistry assays. Gains of chromosomes 7 and 17 without underlying EGFR, HER-2/neu, and TOP2A gene amplification were also observed. A mutation in the K-RAS gene (Gly13Asp GGC>GAC) was identified in the cell line, which was not observed in the six patient samples of apocrine breast carcinomas examined. Similarly, constitutive activation of the MAPK/ERK signaling pathway and deregulation of cell cycle proteins (p16-/pRb-/cyclin D1+ phenotype) with exceedingly high proliferation observed in the MDA-MB-453 cell line were not found in the tissue samples. In conclusion, the MDA-MB-453 cell line shares certain features with apocrine breast carcinoma but differs from patient tissues with regard to various significant characteristics, limiting the value of this cell line as a model for human apocrine breast carcinoma investigations. In contrast to the cell line, EGFR-positive apocrine carcinomas do not harbor K-RAS gene mutations, rendering these tumors amenable to targeted therapy with EGFR inhibitors.

Insulin-like growth factor-II mRNA-binding protein 3 (IMP3) is a member of the insulin-like growth factor-II signaling pathway, and has recently been described as a biomarker of basal-like breast carcinomas. This study explored IMP3 expression in adenoid cystic carcinomas of the breast, a special type of basal-like, triple-negative (estrogen receptor/progesterone receptor/human epidermal growth factor receptor 2/neu protein negative) carcinoma and compared it with a group of apocrine carcinomas, which are an example of estrogen receptor/progesterone receptor negative, special type of breast carcinoma. Eighteen breast adenoid cystic carcinomas (16 primary and 2 corresponding metastases) and 18 apocrine carcinomas (16 invasive and 2 in situ) were evaluated for the expression of IMP3 protein using immunohistochemical method. A cut-off value for IMP3 positivity was set at 10%. Thirteen of 16 (81.3%) primary adenoid cystic carcinomas overexpressed IMP3 protein, predominantly in membranous distribution. The mean percentage of positive cells among primary adenoid cystic carcinomas was 50%. Both metastatic adenoid cystic carcinomas also strongly overexpressed IMP3 protein (70% and 80% of the tumor cells, respectively). In contrast, only 4 of 16 invasive apocrine carcinomas (25%) exhibited IMP3 positivity with significantly lower percentage of positive cells (27%, P<0.001). Two in-situ apocrine carcinomas were negative. Our results indicate that IMP3 may be an additional basal-type marker in breast carcinoma whose expression can be occasionally seen in other types of breast carcinomas such as apocrine type.