Endocervical glandular lesions: a diagnostic approach combining a semi-quantitative scoring method to the expression of CEA, MIB-1 and p16.

OBJECTIVES: To investigate whether combining a semi-quantitative scoring method with the immunohistochemical expression of CEA, MIB-1 and p16, would improve the diagnostic accuracy of endocervical glandular lesions. METHODS: The hematoxylin and eosin-stained sections of 95 cervical biopsies were examined by 4 different observers and were grouped into three categories, benign, dysplasia and adenocarcinoma in situ, depending on the degree of nuclear stratification, nuclear atypia and the number of mitosis and apoptotic figures. Each case was also stained immunohistochemically with antibodies against CEA, Ki-67 (MIB-1) and p16. Staining was graded as negative, weak and positive. The accuracy of the scoring method alone was compared to the accuracy of combining the score with the immunostaining results. RESULTS: Using the semi-quantitative scoring system, most of the cases that were initially diagnosed as atypical hyperplasia or tuboendometrial metaplasia fell into the benign category. This scoring system discriminates effectively (Kruskal-Wallis, p<0.001) between the three categories (benign, endocervical glandular dysplasia and adenocarcinoma in situ). When analyzing the immunohistochemical score, only Ki-67 staining seems to be effective mostly in discriminating between normal glands or glands with atypical hyperplasia and epithelial glandular dysplasia. Ki-67, CEA and p16 failed to discriminate between tuboendometrial metaplasia and epithelial glandular dysplasia. Combining the semi-quantitative scoring system with the immunohistochemical results discriminates between the three categories equally well as the semi-quantitative scoring system alone (Kruskal-Wallis, p<0.001). Nevertheless, the proportion of cases that were classified similarly to the prestudy diagnosis was higher when the combined score was used. CONCLUSIONS: Combining a semi-quantitative scoring scheme with the immunohistochemical expression of CEA, MIB-1 and p16 seems to be of value in classifying some endocervical glandular lesions.

Myoepithelial cell cocktail (p63+SMA) for the evaluation of sclerosing breast lesions.

Sclerosing breast lesions may sometimes mimic the appearance of infiltrating carcinoma due to the entrapment of ductular structures in a fibrotic core. The immunohistochemical detection of the outer myoepithelial cell layer that is indicative of a non-infiltrating lesion is a valuable clue for the diagnosis of such ambiguous cases. The myoepithelial cell markers smooth muscle actin (SMA) and p63 are most commonly used since their specificity and sensitivity are well established. However, recent studies have indicated that some morphologically distinct myoepithelial cells fail to stain for SMA and that p63 positivity can be rarely expressed by a subset of malignant epithelial cells. Moreover, SMA can also be positive in stromal myofibroblastic cells and normal vessels that can be found close to the entrapped ductules and might be erroneously interpreted as myoepithelial cells. In this study, we used a double-immunolabeling technique combining both SMA and p63 antibodies (myoepithelial cell cocktail), in order to investigate whether this technique is advantageous over either marker used alone, in diagnosing sclerosing breast lesions. Our results indicate that p63 alone is not a useful myoepithelial cell marker if applied in large sclerosing breast lesions, however, in smaller lesions it is still of high value. On the contrary, SMA proved significantly useful in the evaluation of myoepithelial cells in larger but not in smaller complex sclerosing lesions. The myoepithelial cell cocktail has a staining sensitivity identical to that of SMA. Nevertheless, in a certain number of cases the cocktail might be useful in differentiating myoepithelial cells from stromal myofibroblasts or vascular smooth muscle cells due to the false impression of a higher staining intensity of the cocktail resulting from the expression of both nuclear and cytoplasmic/membranous antibodies that occupy a wider area of the cell under control.

Cyclooxygenase-2 protein expression in relation to apoptotic potential and its prognostic significance in bladder urothelial carcinoma.

BACKGROUND: Cyclooxygenase-2 (COX-2), a critical enzyme in the conversion of arachidonic acid to prostaglandin E2, influences the biological behavior of human tumors, being involved in carcinogenesis, tumor progression, reduced apoptosis and differentiation. The aim of the present study was to investigate the role of COX-2 protein expression in urothelial carcinoma (UC) of the urinary bladder in relation to clinicopathological data and indices of apoptotic potential. MATERIALS AND METHODS: Immunohistochemistry was applied to 134 paraffin-embedded specimens of UC for the detection of COX-2, p53, bcl-2, caspase-3, bax protein, MLH1 and hTERT. RESULTS: Ninety-four UCs (70.1%) had an enhanced expression of COX-2. The COX-2 semi-quantitative expression was unrelated to tumor grade and local invasion, but it was positively linked with caspase-3 (CPP32) and bax protein semi-quantitative immunoreactivity (p = 0.007 and p = 0.026), as well as with the quantitative expression of MLH1 (p = 0.019). COX-2 was also found to be inversely correlated with the nuclear localization of the catalyst component of the telomerase complex, hTERT (p = 0.009). Multivariate statistical analysis showed that COX-2 immunopositivity was independently associated with worse prognosis of patients with non muscle-invasive UCs (p = 0.002). CONCLUSION: COX-2 overexpression, being possibly a subsequence of apoptosis activation, is associated with an unfavorable overall survival of patients with pTa-T1 UCs.

Matrix metalloproteinase expression in human breast cancer: an immunohistochemical study including correlation with cathepsin D, type IV collagen, laminin, fibronectin, EGFR, c-erbB-2 oncoprotein, p53, steroid receptors status and proliferative indices.

Matrix metalloproteinases (MMPs) are a group of enzymes thought to be responsible for both normal connective tissue matrix remodelling and accelerated breakdown associated with tumour development. The current study aimed to investigate the immunohistochemical expression of matrix metalloproteinase 3 (MMP-3, stromelysin-1) in correlation with the expression of Basement Membrane (BM) antigen (type IV collagen, laminin), fibronectin, cathepsin D, p53, c-erbB-2, proliferative activity (Ki-67, PCNA), steroid receptor content as well as to the other conventional clinicopathological parameters in breast cancer. This study was performed on a series of frozen and paraffin sections from 84 breast cancer specimens by immunohistochemistry using the monoclonal antibody MMP-3 (Ab-1). Stromelysin-1 (ST1) was observed in about 10% of epithelial cells in the control groups (cases of fibrocystic and benign proliferative breast disease), while expression (> 10% of expression) was detected in 89.7% of tumours. The expression of ST1 in carcinoma cells was strongly associated with its presence in the stroma (p < 0.001). A significantly positive correlation was found between ST1 expression, and p53 tumour suppressor gene product (p = 0.004), and a relationship with c-erbB-2 protein and progesterone receptor status was also indicated. These findings suggest that ST1 expression in breast cancer tissue is irrespective of the expression of the extracellular matrix component, the proteolytic enzyme cathepsin D and the growth fraction of the tumour, and that it could be a potential new prognostic marker in breast cancer.

p53/MDM-2 immunohistochemical expression correlated with proliferative activity in different subtypes of human sarcomas: a ten-year follow-up study.

The aim of this study was the evaluation of p53/MDM-2 protein overexpression in different subtypes of human sarcomas, and their correlation with proliferative activity and patient outcome. We selected 40 cases of human sarcomas comprising 6 malignant fibrous histiocytomas (MFH), 1 fibrosarcoma, 1 dermatofibrosarcoma protuberans, 5 liposarcomas, 9 leiomyosarcomas, 1 rhabdomyosarcoma, 3 synovial sarcomas, 2 osteosarcomas, 1 chondrosarcoma, 4 Ewing's sarcomas, 2 Kaposi's sarcomas, 1 malignant haemangiopericytoma, 1 phylloides cystosarcoma, 1 neuroblastoma, 1 chordoma and 1 unclassified sarcoma. All the immunohistochemical markers, which had been used for the characterization of these sarcomas were re-examined. Additionally, the Streptavidin-Biotin peroxidase method was performed on paraffin sections using the monoclonal antibodies: anti-p53 antibody DO7, anti-MDM-2 antibody IF2 and anti-Ki-67 antibody MIB-1. According to our results, p53 protein nuclear expression was detected in 20% (8/40) of the tumours (1 fibrosarcoma, 2 liposarcomas, 1 leiomyosarcoma, 1 rhabdomyosarcoma, 2 Ewing's sarcomas and 1 unclassified sarcoma). MDM-2 nuclear staining was determined in 7.5% (3/40) of the cases (1 MFH and 2 liposarcomas). A high proliferative index was demonstrated in 27.5% (11/40) of the tumours (2 MFH, 4 leiomyosarcomas, 1 rhabdomyosarcoma, 1 osteosarcoma, 2 Ewing's sarcomas and 1 unclassified sarcoma). p53 overexpression was associated with high tumour grade (p < 0.05) and MIB-1 expression was correlated with reduced survival (p < 0.05), but p53 overexpression was not significantly associated with either MIB-1 score or with overall survival of the patients. In conclusion, from this limited and heterogeneous sample of cases, we suggest that the p53/MDM-2 pathway is involved in the tumourigenesis of several sarcoma subtypes, but it is unclear if the overexpression of these genes may become prognostic marker for patients affected with these highly aggressive tumours.

p53 nuclear accumulation in preneoplastic lesions and stage I uterine endometrioid adenocarcinoma.

A total of 118 endometrial neoplastic and preneoplastic lesions comprising 43 uterine endometrioid adenocarcinoma at stage I, 40 complex (adenomatous) hyperplasias and 35 atypical hyperplasias were examined for p53 nuclear accumulation to assess the incidence of p53 alterations in infiltrating carcinomas and to verify if p53 aberrations may allow the identification of a subset of premalignant cases with high risk of progression. No specific immunostaining was observed in the cases of complex hyperplasia without atypias. One (3%) of 35 atypical hyperplasias showed focal areas of p53 immuno-reactivity. The overall frequency of p53 overexpression in endometrial carcinomas was 54%. The distribution of cases with nuclear accumulation of p53 was significantly different (p=0.01) in tumours with different degree of invasiveness. In addition, p53 nuclear accumulation was observed more often in tumours with moderate (G2) or poor differentiation (G3) (p=0.03). Our data indicate that p53 aberrations are not early events in endometrial carcinogenesis and may be related with tumour progression and aggressiveness.

The prognostic significance of epidermal growth factor receptor (EGFR), C-erbB-2, Ki-67 and PCNA expression in breast cancer.

The evaluation of the immunohistochemical expression of epidermal growth factor receptor (EGFR), c-erbB-2 oncoprotein, proliferating indices Ki-67, and PCNA were determined on 68 primary breast carcinomas. These markers were correlated with each other and with other clinicopathological variables such as: age, tumor size? histotype, tumor grade and steroid receptors' content as well as nodal status. The monoclonal antibodies anti-human Epidermal Growth Factor Receptor (EGFR1), anti-human Ki67 (DAKO) and N13259 (Oncogene Science) were applied to paraffin and frozen tissue sections. All markers showed an heterogeneous pattern of staining. There was almost equally high staining intensity at the membranus for EGFR and c-erbB-2 in about 32% of the cases. The EGFR and c-erbB-2 positive cases were much less common in infiltrating lobular (2,2/13) rather than in ductal adenocarcinomas (21,20/55). The low grade carcinomas showed low expression of EGFR and c-erbB-2 oncoprotehl comparing with high grade ductal adenocarcinomas. A high index of Ki-67 was correlated with EGFR and c-erbB-2 membrane positivity. There was an inverse relationship between the expression of c-erbB-2 and EGFR, when compared with oestrogen receptors' content. A significant correlation was also demonstrated between EGFR and c-erbB-2 immunoreactivity with lymph node status. Our results provide evidence that the synchronous immunohistochemical detection of EGFR, c-erbB-2 and Ki-67 may be of useful significance in breast cancer patients, especially when combined with other clinicopathological variables. Furthermore the expression of EGFR and c-erbB-2 oncoprotein may affect the cell proliferation and differentiation.

Prognostic significance of C-erbB-2 and hormone receptors' status in human benign and malignant breast lesions.

In order to investigate the prognostic significance of c-erbB-2, estrogen receptors (ER) and progesterone receptors (PgR), an immunohistochemical staining was performed on 330 tissue sections from paraffin blocks of 50 fibrocystic diseases and 40 ductal adenocarcinomas N.O.S. type, grade II (20 with lymph node metastases (L.N.M)). The positivity for c-erbB-2 was considered only in the cytoplasmic membrane, while for ER and PgR in the nucleus of epithelial cells. All markers showed a heterogenous pattern of staining. Our results imply that benign lesions were negative for c-erbB-2 and hormone receptors except for limited areas with papillary proliferations. Elevated expression of these markers was noted in the adenocarcinomas. No significant difference was observed between the percentage of all markers in primaries and L.N.M. lesions. Hormone receptors' status showed no significant correlation with c-erbB-2 expression. Our results suggest that c-erbB-2, ER and PgR, especially when combined with clinicopathological parameters, may show some prognostic usefulness in breast disease.

Monoclonal gammopathy in chronic myeloproliferative disorders.

The incidence of monoclonal gammopathy in 61 patients with chronic myeloproliferative disorders (CMPD) was studied. The distribution of patients among the CMPD subgroups was: chronic myelocytic leukemia, 24 patients; myelofibrosis, 11; polycythemia vera, 15; essential thrombocythemia, 7; unclassified MPD, 4 patients. Monoclonal gammopathy was found in 5 patients (8.2%). Two of these patients (1 IgA/k and 1 IgM/k) had myelofibrosis and 3 (2 IgG/k and 1 IgG/lambda) polycythemia vera. The presence of monoclonal gammopathy indicates an involvement of the lymphoplasmatic system in CMPD.