Human breast cancer and lymph node metastases express Gb3 and can be targeted by STxB-vectorized chemotherapeutic compounds.

BACKGROUND: The B-subunit of Shiga toxin (STxB) specifically binds to the glycosphingolipid Gb3 that is highly expressed on a number of human tumors and has been shown to target tumor cells in mouse models and ex vivo on primary colon carcinoma specimen. METHODS: Using a novel ex vivo STxB labeling (ESL) method we studied Gb3 expression in cytological specimens of primary human breast tumors from 107 patients, and in synchronous lymph node metastases from 20 patients. Fluorescent STxB was incubated with fine-needle aspiration (FNA) specimens, and Gb3 expression was evaluated by fluorescence microscopy. Furthermore, 11 patient-derived human breast cancer xenografts (HBCx) were evaluated for expression of Gb3 by ESL and FACS. In addition, the biodistribution of fluorescent STxB conjugate was studied after intravenous injection in a Gb3 positive HBCx model. RESULTS: Gb3 expression was detected in 62 of 107 patients (57.9%), mainly in epithelial tumor cells. Gb3 positivity correlated with estrogen receptor expression (p≤0.01), whereas absence of Gb3 expression in primary tumors was correlated with the presence of lymph node metastases (p≤0.03). 65% of lymph node metastases were Gb3 positive and in 40% of tested patients, we observed a statistically significant increase of metastatic Gb3 expression (p≤0.04). Using concordant ESL and flow cytometry analysis, 6 out of 11 HBCx samples were scored positive. Intravenous injections of fluorescent STxB into HBC xenografted mice showed preferential STxB accumulation in epithelial cells and cells with endothelial morphology of the tumor. CONCLUSION: The enhanced expression of Gb3 in primary breast carcinomas and its lymph node metastases indicate that the development of STxB-based therapeutic strategies is of interest in this pathology. Gb3 expressing HBCx can be used as a model for preclinical studies with STxB conjugates. Finally, the ESL technique on FNA represents a rapid and cost effective method for the stratification of patients in future clinical trials.

Loss of ARID1A protein expression occurs as an early event in ovarian clear-cell carcinoma development and frequently coexists with PIK3CA mutations.

ARID1A is a recently identified tumor suppressor gene that is mutated in ∼50% of ovarian clear-cell carcinomas. This mutation is associated with loss of ARID1A protein expression as assessed by immunohistochemistry. The present study aimed at determining the timing of the loss of ARID1A protein expression during the development of ovarian clear-cell carcinoma and assessing its relevance in correlation to PIK3CA gene mutations. A total of 42 clear-cell carcinoma cases with adjacent putative precursor lesions (endometriosis-associated carcinoma cases (n=28) and (clear-cell) adenofibroma-associated carcinoma cases (n=14)) were selected and subjected to immunohistochemical analysis for ARID1A protein expression and direct genomic DNA sequencing of exons 9 and 20 of the PIK3CA gene. ARID1A immunoreactivity was deficient in 17 (61%) of the 28 endometriosis-associated carcinomas and 6 (43%) of the 14 adenofibroma-associated carcinomas. Among the precursor lesions adjacent to the 23 ARID1A-deficient carcinomas, 86% of the non-atypical endometriosis (12 of 14) and 100% of the atypical endometriosis (14 of 14), benign (3 of 3), and borderline (6 of 6) clear-cell adenofibroma components were found to be ARID1A deficient. In contrast, in the 19 patients with ARID1A-intact carcinomas, all of the adjacent precursor lesions retained ARID1A expression regardless of their types and cytological atypia. Analysis of 22 solitary endometrioses and 10 endometrioses distant from ARID1A-deficient carcinomas showed that all of these lesions were diffusely immunoreactive for ARID1A. Among the 42 clear-cell carcinomas, somatic mutations of PIK3CA were detected in 17 (40%) tumors and majority (71%) of these were ARID1A-deficient carcinomas. These results suggest that loss of ARID1A protein expression occurs as a very early event in ovarian clear-cell carcinoma development, similar to the pattern of PIK3CA mutation recently reported by our group, and frequently coexists (not mutually exclusive) with PIK3CA mutations.

Clear cell adenocarcinoma associated with clear cell adenofibromatous components: a subgroup of ovarian clear cell adenocarcinoma with distinct clinicopathologic characteristics.

We occasionally encountered clear cell adenofibromatous (CCAF) components coexisting in the ovarian clear cell adenocarcinoma (CCA). To reveal the clinicopathologic significance of CCAF components in CCA, we classified 67 cases of surgically resected CCA into CCA with and without CCAF components [CCAF(+) and (-) groups], and compared clinicopathologic parameters, that is, patient age, clinical stage, the degree of optimal cytoreduction, patient outcome, histologic grade and Ki-67 labeling index of the CCA, and the presence of endometriosis, between these 2 groups. Fourteen cases (21%) and 53 cases were classified as CCAF(+) and CCAF(-) groups, respectively. Of these 14 CCAF(+) cases, the CCAF components with atypia were observed adjacent to the CCAF components without atypia in 10, and adjacent to the obvious CCAs in 13 cases. In comparison with the CCAF(-) group, the CCAF(+) group showed a higher frequency of histologically low-grade tumors [93% (13 of 14) vs. 43% (23 of 53), P=0.0027], a lower Ki-67 labeling index (mean 35.9% vs. 44.0%, P=0.0492), and better patient prognosis (5-year survival 78.8% vs. 49.3%, P=0.0277). Endometriosis was much less frequent in the CCAF(+) group than in the CCAF(-) group [14.7% (2 of 14) vs. 67.9% (36 of 53), P=0.00096]. Multivariate analysis identified only optimal cytoreduction as independent favorable prognostic factor. These results suggest that CCAF besides endometriosis is associated with the development of CCA, and that the CCAF(+) group may be a distinct subgroup of CCA with less aggressive biologic behavior.

[Solitary fibrous pseudopapillary tumor of the lung: pulmonary fibroadenoma and adenofibroma revisited]. / Tumore fibroso solitario pseudopapillare del polmone: fibroadenoma e adenofibroma polmonari rivisitati.

We describe a peculiar pulmonary lesion, that we interpreted as a pseudopapillary variant of solitary fibrous tumor. The patient was a 62-year-old asymptomatic male, non smoking, presenting with a peripheral nodule, 0.8 cm across, located in the lower lobe of the right lung. The patient is alive and well 18 months after surgical excision of the nodule. Microscopically, the lesion was well-circumscribed and characterized by a diffuse pseudopapillary pattern. Pseudopapillae were large, and were covered by a rim of cubic epithelium devoid of atypia. The stromal axis was fibrous and contained scattered bland spindle cells. Immunohistochemically, the latter were strongly positive for vimentin and CD34, focally positive for BCL2 and CD99, negative for cytokeratin, EMA, TTF1, calretinin, smooth muscle actin, desmin and S100 protein; the epithelial cells were immunoreactive for cytokeratin, EMA and TTF1. We interpret this lesion as a peculiar pseudopapillary variant of solitary fibrous tumor, corresponding to what has been reported in the literature as pulmonary adenofibroma and fibroadenoma. The most important differential diagnostic considerations are briefly discussed.

Biliary adenofibroma: a rare neoplasm of bile duct origin with an indolent behavior.

We report a case of biliary adenofibroma in a 47-year-old woman, who presented with right upper quadrant pain for several months. Abdominal imaging revealed a 16-cm solid and cystic mass in the left hepatic lobe. Histologically, the tumor showed two distinct components: 1) cystic and tubular structures lined by low columnar to cuboidal biliary-type epithelium, and 2) a dense fibrous stroma composed of spindle-shaped cells with only mild nuclear pleomorphism and inconspicuous nucleoli. Mitoses and stromal invasion were absent. The glandular epithelium stained positively for keratin AE.3/Cam 5.2, cytokeratin 7, cytokeratin 19, carcinoembryonic antigen, and epithelial membrane antigen and had a low Ki-67 proliferative index. In addition, the epithelium was positive for D10 but did not stain for 1F6 or acid mucin with alcian blue stain. This staining pattern, similar to bile duct hamartoma (von Meyenburg complex) with which this tumor shares morphologic similarity, suggests that biliary adenofibroma originates from interlobular or larger bile ducts. Three years after a subtotal resection no metastasis or significant tumor growth was noted. However, given the marked nuclear p53 immunoreactivity and tetraploidy status observed in this tumor, we cannot exclude that biliary adenofibroma may represent a premalignant process that warrants complete resection and thorough histopathologic examination.

Adenomyolipoma of the uterus: a case report.

Adenomyolipoma of the uterus is a rare, benign, polypoid lesion considered to be of hamartomatous origin or represent an unusual type of benign Müllerian mixed tumour with a heterologous element. The authors present a case of uterine adenomyolipoma and discuss its pathogenesis. A 62-year-old woman complained of lower abdominal pain and postmenopausal bleeding. Imaging techniques revealed a solid ovarian mass and a polypoid intrauterine lesion. The frozen section diagnosis of the ovarian mass was a thecoma. A total hysterectomy and bilateral salpingo-oophorectomy were performed. On gross examination a pedunculated, polypoid lesion of 7x4.5x3cm was found in the uterine cavity. Microscopically, the polypoid lesion contained both epithelial and mesenchymal elements. The epithelial elements were endometrial glands of various size, formed by proliferative endometrial cells. The mesenchymal elements were composed of endometrial stroma, smooth muscle and mature adipocytes. Both the epithelial and the mesenchymal elements showed a benign appearance, were intermingled with each other and periglandular stromal condensation was absent. The lesion had an irregular surface. Microscopic diagnosis was an adenomyolipoma. The peculiar shape and microscopic features of this lesion suggested that it was a variant of benign Müllerian mixed tumour.

The spectrum of metanephric adenofibroma and related lesions: clinicopathologic study of 25 cases from the National Wilms Tumor Study Group Pathology Center.

The authors report nine new metanephric adenofibroma (MAFs; previously termed nephrogenic adenofibroma) and 16 related tumors from the files of the National Wilms Tumor Study Group Pathology Center (NWTSGPC). All tumors contained a variable amount of a bland spindle cell stroma, which is essentially identical to the recently described metanephric stromal tumor (MST). Features that distinguish this stroma from congenital mesoblastic nephroma (CMN) include intratumoral angiodysplasia, concentric cuffing of entrapped tubules ("onion skinning"), and heterologous differentiation. The epithelial components of these lesions spanned a wide range of appearances. All tumors contained at least focally an inactive embryonal epithelium identical morphologically to metanephric adenoma (MA), and hence each case could be classified as containing MAF. The epithelium of nine tumors had this appearance throughout, and hence these were considered usual MAFs. The epithelium of four tumors demonstrated increased mitotic activity but was otherwise similar to MA. The epithelial component of seven tumors spanned a morphologic spectrum from inactive MA to malignant epithelial predominant Wilms tumor (WT), with gradual transitions noted in several cases. Five other tumors contained a carcinomatous component distinct from these lesions but identical morphologically to papillary renal cell carcinoma (PRCC). In one of these cases, this component had metastasized to the regional lymph nodes at the time of diagnosis. No tumor recurred during follow-up, although almost all patients received adjuvant therapy for WT regardless of their tumor's histology and NWTSGPC diagnosis. In conclusion, MAF is a biphasic tumor that spans the morphologic spectrum between benign pure stromal (MST) and pure epithelial (MA) lesions, and can merge with the morphology of WT, supporting the concept that these are all related lesions. A relationship to PRCC is also evident.

Eccrine syringofibroadenoma (Mascaro): an immunohistochemical study.

Eccrine syringofibroadenoma is an uncommon benign eccrine tumor, which was first described by Mascaro in 1963. It usually develops on the extremities of elderly persons. We report on a 74-year-old man who presented with a 2-year history of a slowly growing lesion on his face. A detailed histologic and immunohistochemical study was performed on the biopsy material. The tumor consisted of epidermal-derived anastomosing thin epithelial cords embedded in a fibrovascular stroma. The epithelial cords contained ductal and cystic structures lined by luminal cells, which were decorated by antibodies against carcinoembryonic antigen, keratin K19, K8, and K18. Antibody to keratin K6 decorated the luminal walls of the acrosyringia. Antibodies to filaggrin decorated the superficial luminal structures. These results suggest dual acrosyringial and dermal duct differentiation in syringofibroadenoma.

Metanephric adenofibroma: report of a case and review of the literature.

The recent recognition of a variety of pediatric renal tumors of different biologic behavior places an ever-increasing demand on the surgical pathologist for an accurate diagnosis. Although metanephric adenofibroma is one of the rarest benign renal tumors, the clinical importance of correctly diagnosing it cannot be overemphasized because it can potentially be mistaken as Wilms' tumor. We describe the clinical, radiologic, and pathologic features of a case of metanephric adenofibroma and discuss its differential diagnosis. The neoplasm was composed of two discrete components: a major fibroblastic element and a minor immature epithelial element. The latter formed a small nodule beneath the renal capsule, which could barely be detected by magnetic resonance imaging. This subcapsular nodule, however, was slightly soft and tan and was distinctly different from the white, whorled cut surface of the main tumor. It was formed by closely packed small immature epithelial cells in a slightly edematous background, which was histologically identical to metanephric adenoma and closely resembled epithelial Wilms' tumor. Unlike Wilms' tumor, however, the epithelial cells were very bland with no mitoses. The main bulk of the tumor was formed by spindle fibroblastic cells that were cytologically similar to the spindle cells in congenital mesoblastic nephroma. The tumor, however, was well demarcated without the irregular infiltrating edges of congenital mesoblastic nephroma. In contrast to the randomly distributed epithelial element throughout the stromal component in previous reported cases of metanephric adenofibroma, our finding of the exceedingly small and discrete epithelial component expands the known histologic spectrum of the disease. In addition, the presence of such minute epithelial nodule underscores the importance of diligent pathologic examination and careful sampling of tissue for histologic examination.

Papillary and metanephric adenomas of the kidney.

Papillary renal adenomas are the commonest neoplasms of the renal tubular epithelium, occurring in up to 40% of adults. If these criteria are met, the diagnosis of adenoma can be made with confidence: (1) papillary, tubular, or tubulopapillary architecture; (2) diameter less than or equal to 5 mm; and (3) does not histologically resemble clear cell, chromophobe, or collecting duct renal cell carcinomas. They are composed of epithelial cells with variable amounts of cytoplasm with variable tinctorial qualities including clear. Metanephric adenoma and metanephric adenofibroma are closely related neoplasms that appear to be benign. Although they have a morphological resemblance to Wilms' tumor, there is some genetic evidence relating them to papillary adenoma and papillary renal cell carcinoma. Metanephric adenoma and metanephric adenofibroma occur at all ages, have a 2:1 predominance of female patients, and are associated with polycythemia. No instance of malignant behavior has been reported. The adenomatous component is composed of uniform small cells arranged in tubular or papillary architectural patterns. The fibromatous component is moderately cellular and composed of bland spindle cells.

E-cadherin expression in human epithelial ovarian cancer and normal ovary.

The ovarian surface epithelium (OSE) is the origin of the majority of human ovarian cancers. These adenocarcinomas are characterized by initial local growth followed by spreading into the peritoneal cavity at later stages of tumor progression. The cell-adhesion molecule E-cadherin (E-cad) plays an important role in maintaining tissue integrity. Disappearance or impaired function of E-cad have often been associated with tumor formation and invasion in vivo and in vitro. The cell-specific expression of E-cad was investigated in normal human ovaries (n = 12), in benign (n = 5) and borderline (n = 4) ovarian epithelial tumors and in adenocarcinomas of different stages and histological grades (n = 18), by immunohistochemistry and immunoblotting. An ovarian cancer cell line (NIH-OVCAR3) was used as a reference. The epithelial origin of the cells was confirmed with cytokeratin (AE1/AE3) staining. In normal ovaries, the expression of E-cad was limited to inclusion cysts or deep clefts lined with OSE, whereas no staining of the OSE could be demonstrated at the surface of the ovary. In contrast, benign and borderline tumors uniformly expressed E-cad. This was observed in malignant tumors of all stages despite their degree of differentiation. E-cad was also present in metastasis from such tumors. The cell-specific expression of E-cad in inclusion cysts of normal ovaries and in epithelial layers of borderline tumors indicates a role for E-cad in the early events of the progression to a malignant phenotype. E-cad was not downregulated in later stages of ovarian cancer progression.

p53 mutation in epithelial ovarian carcinoma and borderline ovarian tumor.

We have investigated a series of 19 human ovarian carcinomas and 17 borderline ovarian tumors to determine the loss of heterozygosity on chromosome 17p and possible concurrent p53 mutations. Allelic losses were assessed using restriction fragment length polymorphism study, and p53 gene mutations were detected by single strand conformation polymorphism analysis and by direct sequencing. In addition, we stained the same tumor sections immunohistochemically to detect p53 protein in tissues. Among 19 ovarian malignant tumor samples tested, we identified 17p allelic deletions in 12 (63.2%) of 19 informative cases. The p53 gene mutation was observed in 7 of 19 (36.8%) malignant ovarian tumors, and it was predominantly observed in tumors with allelic loss on 17p (six of seven tumors, 85.7%). Although 9 cases of 17 borderline ovarian tumors showed shifted bands on single strand conformation polymorphism analysis, only one case was proved to have a point mutation in direct sequencing. We also obtained six cases (31.6%) of positive immunoreactivity from 19 ovarian cancers and 3 cases (17.6%) from 17 borderline ovarian tumors. We conclude that loss or inactivation of tumor suppressor gene function by chromosome 17p allelic deletions or p53 mutations are important genetic changes in ovarian cancer.

Histologic grade and cellular proliferation at the deepest invasive portion correlate with the high malignancy of submucosal invasive gastric carcinoma.

To establish appropriate indications for endoscopic treatment of submucosal invasive gastric carcinoma, we investigated clinicopathologic features and proliferating cell nuclear antigen (PCNA) expression at the deepest invasive portion of 192 differentiated submucosal invasive gastric carcinomas that had been surgically resected. Lymph node metastasis was demonstrated in 30 (15.6%) of 192 lesions. Histologic heterogeneity (based on differentiation at the deepest invasive portion) was demonstrated in 36 (18.8%) of the 192 lesions. In 159 lesions, excluding 33 undifferentiated lesions at the deepest invasive portion, the depth of invasion, histologic grade, lymph vessel involvement, infiltrative growth pattern (INF) and existence of an ulceration were all significantly correlated with the incidence of lymph node metastasis. The lesions with both well-differentiated adenocarcinoma (WELL) and minimal submucosal invasion (sm1) showed no lymph node metastasis. PCNA expression was estimated in 59 good stained lesions. The mean PCNA labeling index (LI) was 50.9 +/- 7.2% in lesions with lymph node metastasis and 43.7 +/- 9.3% in those without lymph node metastasis (p < 0.05). In addition, PCNA-LI also correlated significantly with the histologic grade, depth of invasion, INF and lymph vessel involvement. These results indicate that the submucosal invasive gastric carcinoma with both WELL and sm1, which shows no other risk factors, can be considered as the appropriate indication for curative endoscopic treatment. The PCNA-LI at the deepest invasive portion is useful in understanding the biology of lymph node metastasis in submucosal invasive gastric carcinoma.

Expression of opioid peptides in cells and stroma of human breast cancer and adenofibromas.

The expression of beta-endorphin, Met-enkephalin and Leu-enkephalin was studied in 63 malignant and benign human breast tumors using immunohistochemical methods. Among invasive ductal carcinomas, 93% were positive for beta-endorphin, 87% for Leu-enkephalin and 90% for Met-enkephalin, in both the tumor stroma and the cell bodies. Enkephalin was predominant in cells, whereas endorphin was predominant in stroma. Nearly the same distribution was found in adenofibromas. In pericancerous normal tissue, neuropeptides were predominantly expressed in the stroma. Although the neuropeptide expression is not cancer-specific, it could be cancer-related, since the results suggest that the neuropeptide expression could reflect the host response to cancer cells and not only the cancer cell activity. The possibility of a direct action of the nervous system on stroma reaction and then on cancer cells is discussed.

Vimentin expression in benign and malignant breast epithelium.

AIMS: To determine vimentin expression in epithelial cells in benign breast disease and malignant breast tumours; to assess the value of vimentin expression as a prognostic indicator in breast carcinoma. METHODS: Frozen and formalin fixed, paraffin wax embedded sections from 78 carcinomas, three phyllodes tumours, 19 fibroadenomas and 19 cases of fibrocystic disease were examined with a monoclonal antibody from the V9 clone. A correlation between vimentin expression and known prognostic indicators was sought in ductal carcinomas. The intracellular localisation of vimentin was examined in benign and malignant lesions. RESULTS: Vimentin expression was identified on frozen section in the cells of ductal (53%), lobular (86%), and mucinous (33%) carcinomas and in the luminal epithelium of fibroadenomas (68%), cases of fibrocystic disease (47%), and a malignant phyllodes tumour. Formalin fixation reduced the percentage of carcinomas and cases of benign disease in which vimentin was detected. This reduction was more pronounced in fibroadenoma and fibrocystic disease than in ductal carcinoma. Associations were identified between vimentin expression as detected on frozen section and tumour grade, size, number of lymph nodes affected, oestrogen receptor content and growth fraction. Only the association with grade was significant (p = 0.045). There was no significant correlation between any of these prognostic variables and vimentin expression on paraffin wax sections. There was no difference in the intracellular localisation of vimentin staining between benign and malignant lesions, or between low and high grade ductal carcinomas. CONCLUSION: There is some loss of vimentin immunoreactivity after formalin fixation. Vimentin expression does not assist in differentiating between benign and malignant breast disease, but is correlated with tumour grade in ductal carcinoma.

Biliary adenofibroma. A heretofore unrecognized benign biliary tumor of the liver.

Benign biliary tumors of the liver are uncommon. In this report, we describe a distinctive biliary tumor of 7-cm diameter occurring in the right lobe of the liver of a 74-year-old Chinese woman. The lesion, characterized by a complex tubulocystic nonmucin secreting biliary epithelial and an abundant fibroblastic stromal components, is distinct from other well-recognized biliary lesions. A number of unusual features are focally present, namely, intraluminal bile concretions, apocrine-like epithelial change, acute inflammation, and granuloma. The tumor shows a striking resemblance to Meyenburg's complex (MC), but the large size of the lesion and the absence of any typical MC in the background liver are exceptional for the latter. Its expansile growth, possession of mitoses, and foci of epithelial tufting and cellular atypia favor a neoplastic process. Previous reported cases of adenomatous neoplastic transformation of MC are dissimilar. We therefore conclude that this is a hitherto unrecognized biliary tumor that may be yet another neoplastic form of MC and propose the designation biliary adenofibroma. The course appears benign, but malignant epithelial transformation may supervene if the lesion is left untreated.

Abundant expression of immunoreactive endothelin 1 in mammary phyllodes tumor: possible paracrine role of endothelin 1 in the growth of stromal cells in phyllodes tumor.

Immunoreactive endothelin 1 (irET-1) concentrations were measured in extracts prepared from 4 phyllodes tumors and 14 fibroadenomas. irET-1 was detectable in all tissue extracts by specific radioimmunoassay, and the mean concentration of irET-1 was 18-fold and 27-fold higher in tissue extracts from phyllodes tumors than in those from intracanalicular fibroadenomas and pericanalicular fibroadenomas, respectively. Reverse-phase high-performance liquid chromatography coupled with radioimmunoassay in the extracts from phyllodes tumors revealed one major irET-1 component corresponding to human standard ET-1. Furthermore, immunocytochemical staining for ET-1 revealed that numerous ET-1-immunoreactive cells were seen in the epithelial cells but not in the stromal cells, suggesting that ET-1 is synthesized by the epithelial component of phyllodes tumors. A possible paracrine role of ET-1 in the growth of this rare tumor which is characterized by its prominent stromal cellularity is discussed.