The value of expression of EGFR, telomerase and topoisomerase IIα in malignant effusion smears before and after chemotherapy.

PURPOSE: To evaluate the significance of expression of epidermal growth factor receptor (EGFR), telomerase and topoisomerase IIα (topo IIα) in cells of malignant effusions of patients under chemotherapy. METHODS: We studied the expression of EGFR, telomerase and topo IIα in malignant effusion smears of 95 cancer patients before and after chemotherapy. Immunocytochemical and in situ hybridization techniques were applied. RESULTS: Positive expression before chemotherapy of telomerase, topo IIα and EGFR was found in 64.2, 63.2 and 69.5% of the cases, respectively; the expression of these markers following chemotherapy was 43.6, 28.2 and 53.8%, respectively. The stronger prognostic factor affecting survival before chemotherapy was telomerase (p=0.0002), whereas after chemotherapy the strongest factor was EGFR (p<0.0001). A positivity for all three markers following chemotherapy was associated with shorter survival compared with positivity for only 1 or 2 markers (p<0.0001) or with a negative expression. CONCLUSION: It seems that expression of EGFR, telomerase and topo IIα in malignant effusion smears is adversely affecting prognosis and survival.

Additional slides from residual ThinPrep Pap tests: Of potential diagnostic benefit in equivocal cases?

The aim of this study was to further evaluate the diagnostic significance of additional slides prepared from residual ThinPrep (TP) Pap Tests. Up to 10 repeat slides were prepared from 105 residual TP cervical samples. All additional slides were evaluated for the presence of diagnostic elements which were not found on the primary TP slide. After the evaluation of the repeat slides, an upgraded diagnosis was noted in 15 cases (14.3%). The reclassified cases included: three negative cases reclassified as two ASC-US and as one LSIL, seven cases of ASC-US reclassified as six LSIL and as one HSIL, and five cases of LSIL reclassified as HSIL. The highest rate (7/15 cases, 46.7%) of cases with an upgraded diagnosis was noted in the ASC-US diagnostic category. Our results suggest that repeat processing of residual TP cervical samples may represent an adjunctive diagnostic tool for a more accurate classification of ASC-US cases. Nevertheless, the practical value of this approach seems to be limited by its significant cost and its uncertain effectiveness.

Repeat processing of residual ThinPrep Pap tests: sampling of the vial may not be invariably homogeneous.

OBJECTIVE: To re-evaluate the reproducibility of additional slides prepared from residual cervical ThinPrep (TP) samples. STUDY DESIGN: Sixty paired specimens (conventional smears and direct-to-vial TP) were studied. Up to 10 additional slides were prepared from each TP vial. All slides were reviewed for adequacy of material, presence of abnormal cells and presence of normal flora or other pathogens. The additional TP slides were further evaluated for the presence of diagnostic elements which were not found on the conventional smear and primary TP slide. RESULTS: Abnormal cells found on the primary TP slide were also identified on all additional slides in 48/50 cases (96%) with squamous cell lesions. The distribution of material on TP slides was evaluated as homogenous in 51 cases (85%) and as non-homogenous in 9 (15%). Using the primary slides (conventional smear and TP) as a reference, additional diagnostic cells upgrading the cytologic diagnosis were found on the repeat slides in 7 cases (11.7%) and fungi consistent with Candida in 3 (5%). CONCLUSION: Repeat processing of residual cervical TP samples may not be an invariably reproducible procedure and the first slide may not be necessarily representative of the specimen as a whole. Nevertheless, both primary and repeat TP slides seem to be extremely effective in detecting a lesion (regardless of grade) in abnormal cases. The exact impact of non-homogeneous sampling of the vial on the diagnostic accuracy of the TP method should be further investigated.

Prognostic value of DNA analysis of prostate adenocarcinoma: correlation to clinicopathologic predictors.

The ability to accurately predict tumor behavior and patient survival is a problem in managing patients with prostate cancer. DNA ploidy provides important information for the evaluation of the prognosis of prostate cancer. The aim of this study was to investigate the DNA ploidy in imprints from prostate adenocarcinomas in a group of 70 patients in relation to Gleason score, tumor differentiation, stage and PSA serum levels. The DNA content was studied in Feulgen-stained imprint smears through the image analysis technique using a SAMBA 2005 Image analyzer. According to our measurements, a strong correlation was observed between DNA ploidy status and tumor differentiation (p<0.001). A statistically significant difference was found between DNA aneuploidy and increased pretreatment PSA serum levels (>4 ng/ml) (p<0.001), as well as between ploidy pattern and stage of the disease (p<0.001). Our results conclude that DNA ploidy status appears to be an additional marker in the field of prognosis of prostatic adenocarcinoma and could provide useful information on the potential behavior of prostate cancer.

Expression of p120, Ki-67 and PCNA as proliferation biomarkers in imprint smears of prostate carcinoma and their prognostic value.

The cell proliferation markers p120, Ki-67 and proliferating cell nuclear antigen (PCNA) recognize nuclear antigens. The expression of these proteins by immunostaining methods was reported to be of value in determining the prognosis of patients with malignant diseases. In this study, we evaluated the prognostic significance of the expression of nuclear antigens p120, PCNA and Ki-67 in prostate cancer and compared the results with other prognostic factors. Imprint smear samples obtained from 70 patients immediately after radical prostatectomy for prostatic carcinoma were immunostained with monoclonal antibodies against p120, Ki-67 and PCNA. The immunostaining results were correlated with Gleason score, tumour differentiation, stage and prostatic specific antigen (PSA) levels. Our findings demonstrate that p120, Ki-67 and PCNA expression in prostatic carcinoma smears, correlated significantly with the degree of Gleason score (P < 0.001). When combining p120, Ki-67 and PCNA positivity with tumour differentiation there was a significant association among these parameters (P < 0.001). Overexpression of p120, Ki-67 and PCNA, was also associated with increased PSA serum levels (>4 ng/ml) (P < 0.001). The distribution of p120, Ki-67 and PCNA expression in prostate carcinomas was not statistically significant for Ki-67 (P = 0.69) and p120 (P = 0.22) but was significant for PCNA (P < 0.001) as far as the histological stage (T2a, T2b, T2c, T3a). P120, Ki-67 and PCNA expression had significant prognostic value for disease-free survival. Our results conclude that nuclear antigens p120, Ki-67 and PCNA appear to be additional markers in the field of prognosis of prostatic carcinoma.

Telomerase expression as a marker in prostate cancer: correlation to clinicopathologic predictors.

The aim of this study was to investigate by an in situ hybridization procedure the Telomerase expression as a marker in prostate cancer and to correlate these results with several prognostic factors concerning this cancer. Imprint smear samples were obtained from 70 prostates removed from patients who underwent radical prostatectomy for prostate adenocarcinoma. Telomerase expression in cancerous prostate smears was studied using an in situ hybridization procedure. The results were correlated with prognostic factors such as pathologic staging, Gleason grading, PSA serum levels and tumour differentiation. Positive Telomerase expression was detected in 88.6% prostate cancer smears. Telomerase expression was significantly correlated with the Gleason score (p < 0.001), tumour differentiation (p < 0.001) and PSA serum levels (p = 0.002). The distribution of Telomerase expression according to histopathological staging was not statistically significant (p < 0.56). In conclusion Telomerase expression could be a marker indicating the malignant potential of prostate cancer.

Diagnostic approach of effusion cytology using computerized image analysis.

The objective of this study is to investigate whether image cytometry is a sensitive and specific method for the differential diagnosis of equivocal cells in routine cytology of effusion smears. One hundred four effusion smears were studied from routine cytologic material. Cytologically 56 (53.8%) of the smears were classified as malignant, 26 (24%) as suspicious and 22 (21.1%) as benign. Two morphometric variables (nuclear major axis length and nuclear area) of the nuclei were measured by an image analysis system. Higher values for the area were found for malignant rather than benign and suspicious cells (p < 0.0005 and p < 0.005 respectively). The same result was extracted for the nuclear major axis length values (p < 0.0005 and p < 0.0005 respectively). Values of nuclear major axis length and nuclear area didn't differ significantly between benign and suspicious cells (p = 0.071 and p = 0.066 respectively). The results show that the range of the values for suspicious cells is closer to the range of the benign cells. Cytomorphometry of the effusion smear cells may provide important information for the differentiation of atypical mesothelial cells from malignant adenocarcinoma cells.

Correlations between nuclear/cytoplasmic area ratio and classification of cervical smears.

The aim of this study was to assess the nuclear/cytoplasmic (N/C) ratio using computerized image analysis of cervical smears with intraepithelial neoplasia (CIN) grade I to III associated or not with cellular changes of human papillomavirus (HPV) in an attempt to determine if this method is more sensitive for the estimation of the grade of CIN. One hundred and ten cervical smears from women with a mean age 35.03 years were studied. The cytological diagnosis was as follows: CIN I + HPV (11), CIN II + HPV (11), CIN II + HPV (8), CIN I (7), CIN II (6), CIN III (8), Ca (22), HPV (32), CIN I-II + HPV (2) and CIN II-III + HPV (3). All cases were histologically examined: 93 cases were in agreement and 17 were under- or overestimated cytologically. The morphometric study of cervical smears was carried out by image analysis. Data were analysed by one way analysis of variance followed by Bonferroni test of multiple comparisons. Statistically significant differences were detected between the three grades of CIN or CIN HPV or only HPV (p<0.0001). The results demonstrated that the N/C ratio measured by image analysis on precancerous lesions of cervical smears could be considered as an additional tool for the classification of cervical smears, especially in determining the discrepancies between cytological and histological diagnoses.

Moc-31, fibronectin and CEA in the differential diagnosis of malignant effusions: an immunocytochemical study.

In discriminating benign and malignant origins of cytologically suspicious effusion smears a panel of antibodies against carcinoembryonic antigen (CEA), Fibronectin (F) and MOC-31 was used with immuno-cytochemical techniques. One hundred and thirty seven effusions were studied of which 107 had a malignant and 30 a benign aetiology as determined by clinical and histological examination. Cytologically 24 were diagnosed as benign, 97 as malignant and 14 as suspicious. Staining for F was positive in all effusions of benign and 3 of malignant origin. MOC-31 was positive in 95 (88.8%) of effusions of malignant origin but none of benign origin. Positive CEA was observed in 43% of effusions of malignant origin and in 10 of benign origin. The combination of MOC-31 positivity measured the sensitivity and specificity of the cytological examination in cases where the cytological examination result was suspicious as did F positivity improve the sensitivity for a benign origin of the effusion. Positivity or negativity for CEA is less valuable than the other parameters.

Prognostic significance of p53, bcl-2 and EGFR in carcinoma of the endometrium.

OBJECTIVE: To evaluate the part played by several parameters in the prognosis of patients with endometrial carcinoma. STUDY DESIGN: Eighty imprint smears from fresh endometrial tumor specimens were studied immunocytochemically for the expression of p53, bcl-2 and epidermal growth factor receptor. Also, the presence of estrogen receptor (ER) and progesterone receptor (PR) in the tumor tissue was measured. The data obtained were related to survival, and associations were sought between the parameters studied. RESULTS: Strong associations were found between advanced stage, high grade, lymph node metastases at diagnosis, nonendometrioid histology and p53 expression with poor survival. Bcl-2 expression was associated with good five-year survival. ER expression was associated marginally with good five-year survival, but PR expression was not. A strong association was found between p53 and advanced disease, stage and lymph node metastases at diagnosis. An association between EGFR positivity and survival was not found. CONCLUSION: p53 Expression of uterine tumors is an independent and strong indicator of poor prognosis. Even patients with stage I and II disease at surgery who have p53-positive tumors must be considered at high risk.

p53 and bcl-2 protein expression in non-small-cell lung carcinoma.

The diagnostic significance of p53 and bcl-2 proteins in epithelial non-small-cell lung cancers was examined, and the relationship between these proteins expression and other disease parameters, including stage of the disease and tumor differentiation, were studied. We analyzed p53 and bcl-2 proteins expression in 60 imprint smears of freshly resected lung tumors (37 squamous and 23 adenocarcinomas) using the immunocytochemical technique. There were seven patients with stage I disease, 24 with stage II, 23 with stage IIIa, and six with stage IIIb disease, according to the International Staging System classification. Sixteen of the tumors were bcl-2 positive and 25 were p53 positive. Twenty tumors were negative for both bcl-2 and p53 (33.3%). Statistical analysis showed no association between the incidence of p53 or bcl-2 positivity. Adenocarcinoma or squamous carcinoma analysis showed significant associations between p53 positivity and poor differentiation and advanced disease stage as well as bcl-2 and early disease stage and well-differentiated tumors. There was also an association between the stage of the disease and the degree of differentiation of the tumors. In conclusion, bcl-2 positivity must be considered a good prognostic sign. On the other hand, p53 positivity seems to indicate, even in tumors at a relatively early stage, that a serious aggressive tumor which will not be easily eradicated is present.

Expression of p53, bcl-2 and heat shock protein (hsp72) in malignant and benign ovarian tumours.

The occurrence of p53, bcl-2 and heat shock protein (HSP) expression in ovarian tumours was examined and the correlation was investigated between the expression of these proteins and other disease parameters, including FIGO stage, histological subtype, tumour differentiation and steroid hormone receptor status. We analysed p53, bcl-2 and HSP expression in 100 smears of patients with epithelial ovarian carcinomas, 16 smears of patients with borderline malignancy and 20 smears of patients with benign ovarian neoplasms by using immunocytochemical techniques. There were 29 patients with stage I disease, 24 with stage II disease, 40 with stage III disease and seven with stage IV disease according to the FIGO classification. The sensitivities and specificities of bcl-2, p53 and HSP for malignancy were 53% and 40%, 43% and 80%, and 37% and 90%, respectively. HSP was statistically significantly associated with malignant rather than benign tumours. Significant association was also observed between bcl-2 and p53, and p53 and HSP. The association of HSP with malignant tumours is confined to the premenopausal group of patients and in this group by itself there is also a significant association between p53 and malignancy. HSP and p53 were associated with undifferentiated carcinomas, bcl-2 and p53 expression is reduced as disease stage progresses in serous carcinomas and bcl-2 expression is increased as disease progresses in endometrioid carcinomas. There was no significant association between bcl-2 and ER/PR status. In conclusion, HSP has a high specificity for malignant ovarian tumours, bcl-2 and p53 have only moderate to low sensitivity and specificity. Changes in the frequency of bcl-2 and p53 overexpression between FIGO I and FIGO III stage disease of different ovarian carcinomas indicate a different role of these substances in cellular survival mechanisms in different carcinomas. bcl-2 probably is associated with cell proliferation but not with differentiation.

Expression of cathepsin D and pS2 in imprint smears of breast carcinoma.

The aim of this study was to add to existing information on the effects of certain tumour markers expressed by breast cancers on tumour malignancy as evidenced by size of primary and occurrence of lymph node invasion. One hundred freshly resected breast cancers were examined by immunocytochemical staining of imprint smears for Cathepsin D and pS2. Oestrogen receptor (ER) and progesterone receptor (PR) were tested for by dextrose-coated charcoal (DCC) assay and the results correlated with tumour size, histology and presence or absence of lymph node metastases at the time of surgery using chi(2) analysis. A significant positive correlation was demonstrated between Cathepsin D positivity and ER, PR and pS2 positivity. In tumours < 2 cm in diameter at surgery a positive correlation was observed between Cathepsin D positivity and the presence of lymph node metastases. The findings support the hypothesis that Cathepsin D may promote early metastasis, possibly by its proteolytic activity.

Cathepsin D immunoreactivity in ovarian cancer: correlation with prognostic factors.

In view of the somewhat inconclusive nature of the reports of the role of Cathepsin D (Cath D) in ovarian carcinoma and its relationship with various other parameters of malignancy the present study was performed to aid in the further clarification of this role. One hundred freshly resected primary ovarian carcinomas of various histological types were studied for ER, PR and Cath D status and the results examined with respect to menopausal status, histology, size and lymph node invasion. In our series Cath D positivity was more frequent in serous than in other types of ovarian cancer but this Cath D positivity was not related to the frequency of lymph node invasion regardless of the size of the tumor. Nor was any association observed between Cath D positivity and ER or PR status of the tumors or the menopausal state of the patients. The reported prognostic value of Cath D, ER and PR is discussed as well as the distinction between tumor invasion by lymphatic channels and direct interstitial infiltration. It was concluded that Cath D may not play a role in the former mode but, as might be expected from its proteolytic properties, in local spread by means of tissue destruction.

Primary malignant melanoma of the conjunctiva of the upper eyelid. A case report.

BACKGROUND: Malignant melanoma of the conjunctiva is rare. The nomenclature and clinical and pathologic features of cutaneous and conjunctival melanomas are different. CASE: A 62-year-old male presented with a history of slight bleeding of the upper conjunctiva for the previous six months. On clinical examination the ophthalmologist observed a smooth, partly nodular, pigmented lesion on the conjunctiva under the left eyelid, 1.5 cm in diameter. Fine needle aspiration (FNA) biopsy of the mass showed tumor cells dispersed as single cells with eccentric, round nuclei; coarsely granular chromatin; prominent nucleoli; and dense cytoplasm with occasional brownish pigmentation as well as small aggregates of spindle-shaped neoplastic cells with hyperchromatic nuclei and no cytoplasmic pigment. CONCLUSION: FNA cytology is a simple and efficient method of making the diagnosis of malignant melanoma in conjunctival masses. Careful correlation with the clinical history and histologic findings is often necessary for confirmation of the diagnosis.

Vimentin and alpha-fetoprotein expression in breast cancer smears: relationship with various prognostic factors.

Estrogen (ER) and progesterone (PR) receptor status, factors known to influence the prognosis and therapeutic possibilities in patients with breast cancer; alpha-fetoprotein (AFP), which is an estrogen binder; and vimentin (V) were determined in cytological imprint smears of 75 mastectomy specimens. The results were classified according to tumor type and menopausal state and analyzed to determine possible relationships among these four markers under the different circumstances. There was a positive association of ER positivity with PR positivity, AFP positivity, and V negativity using chi 2 analysis. The lobular carcinomas studied were predominantly ER+, AFP+, V-, while in the ductal carcinomas V positivity was marginally predominant and also frequently associated with ER+ or PR+ status. The small number of medullary carcinomas made statistical analysis of this group difficult. It seemed probable that the marker status of the tumors was dependent more on the tumor type than on the menopausal state of the patients, thorough follow-up, especially of V+ medullary carcinomas, may provide insight into the value of vimentin as a primary rather than a secondary prognostic factor.

Immunocytochemical detection of P-glycoprotein in the management of malignant effusions.

P-glycoprotein (P-gp), a cell membrane protein, has been found in multidrug-resistant cancer cells. A total of 104 smears from patients with breast-cancer-associated pleural effusions and ovarian-cancer-related peritoneal effusions were studied for P-gp with the antibody C-219 and the avidin-biotin-immunoperoxidase method. Samples were taken before and 3 and 7 days after intracavitary bleomycin therapy and reaccumulation of effusion was assessed at 30 days. Smears that were P-gp-negative by the 7th day were associated with a good 30-day response to bleomycin in the majority of cases, while P-gp-positive smears were associated with a significant reaccumulation of fluid at 30 days. P-gp status is a valuable prognostic indicator of response to intracavitary bleomycin treatment in effusions from breast or ovarian cancer.

Immunocytochemical localization of Cathepsin D and CA 125 in ovarian cancer.

OBJECTIVE: To study the expression of Cathepsin D (Cath D) and CA 125 antigens and ER and PR receptors on freshly obtained surgical specimens of ovarian carcinomas and their relationship with menopausal status, tumor histology, primary tumor size and lymph node invasion. METHODS: The tumors obtained from 100 women were measured and cut in half. The cut surface of one half was pressed against glass slides which were air dried and stained using the Avidin-Biotin peroxidase method for Cath D and CA 125 antigens. The slides were viewed under the light microscope for the characteristic brown granules in the cytoplasm or membrane of the malignant cells. The other half of the tumor was subjected to routine histological examination and part used for the demonstration of ER and PR receptors. The results were analyzed using chi 2 analysis. RESULTS: Cath D positivity was as common as CA 125 positivity. Cath D positivity is more frequently associated with serous carcinomas than with others. No relationship was observed between ER/PR positivity and Cath D or CA 125 positivity. CONCLUSION: The high incidence of Cath D positivity makes it a possible complementary method for following up ovarian carcinoma patients especially those who are CA 125 negative.

Expression of cathepsin-D, CA125 and epidermal growth factor receptor in imprint smears of ovarian carcinoma.

One hundred primary ovarian cancers were studied for the expression of cathepsin-D, CA125 and epidermal growth factor receptor (EGF-R) by staining of imprint smears from freshly obtained surgical specimens. Estrogen and progesterone receptors of the primary tumor, lymph node invasion, menopausal status of the patients and primary tumor size were also noted. The polymorphism of the antigenic characteristics of ovarian carcinomas was noted and significant associations of EGF-R positivity and lymph node negativity, EGF-R positivity and serous carcinomas, EGF-R positivity and cathepsin-D positivity, cathepsin-D positivity and CA125 positivity, and CA125 positivity and serous carcinomas were observed. The high incidence of cathepsin-D positivity makes it a possible complementary method to CA125 for following up patients. It is suggested that complete antigenic profiles of individual tumors are more likely to provide accurate prognostic information in individual cases.