In vitro growth characteristics and chemosensitivities of endometrial cancer using a soft agar clonogenic assay

A soft agar clonogenic assay was used to analyze 91 fresh human endometrial tumor samples for in vitro growth and chemosensitivities. Overall cloning success was 81%. Of the 75 samples adequate for drug testing (30 or more colonies per plate), histologic analysis demonstrated 36 adenocarcinomas, seven adenoacanthomas, ten adenosquamous carcinomas, nine mixed mesodermal sarcomas, seven carcinosarcomas, three papillary adenocarcinomas, and three clear-cell carcinomas. Mixed mesodermal sarcoma specimens demonstrated the most efficient growth, followed by the adenosquamous adenocarcinoma and adenoacanthoma samples. In vitro chemosensitivities were determined by colony inhibition resulting from continuous exposure of tumor cells to known achievable peak plasma and one-tenth peak plasma concentrations of a particular agent. The plating efficiency reflected the clinical virulence by cell type. Thus, we found that malignant endometrial tumors can be cloned in vitro, and the observed chemosensitivities reflect the relative clinical resistance of the tumors to these agents. Based on the high cloning success, this tumor type may be well suited as a model using in vitro soft agar cloning for new drug screening, evaluation of novel drug combinations, and studies of human tumor biology.     

A clinicopathologic study of endometrial carcinomas with argyrophil cells.

Of 41 endometrial carcinomas examined with Grimelius staining, 9 tumors were found to be composed predominantly or partially of argyrophil cells. They were 4 well-differentiated adenocarcinomas, 4 moderately differentiated adenocarcinomas, and 1 adenosquamous carcinoma. Argyrophil granules were found mainly in the apical portion and sometimes in the entire cytoplasm of glandular tumor cells in the well- and moderately differentiated adenocarcinomas. In the adenosquamous carcinoma, argyrophil granules were located in the squamous cells as well as in the grandular cells. The distribution of argyrophil granules was in parallel with that of secretory granules identified by electron microscopy. A clinicopathologic study of these 9 cases revealed that the patients with endometrial argyrophil cell carcinoma tended to be associated more frequently with obesity, hypertension, and diabetes mellitus than the patients with usual endometrial carcinoma.     

Prognostic significance of squamous differentiation in stage I endometrial adenocarcinoma

The prognostic implication of benign and malignant squamous differentiation was examined in 267 consecutive patients with stage I endometrial carcinoma. Patients with adenosquamous carcinoma had a significantly poorer ten-year survival rate (54.7%) than patients with adenocarcinoma (70.5%) or adenoacanthoma (87.4%). This was related to a tendency for adenosquamous carcinoma to be associated with poorly differentiated glandular elements and to deeply invade the myometrium. The mean depth of myometrial penetration was 57% for adenosquamous carcinoma compared with 24% for adenocarcinoma and 19% for adenoacanthoma. To examine the prognostic significance of malignant squamous differentiation independently of the grade of the associated glandular component, the subgroup of patients with well-differentiated adenocarcinoma was compared. Patients with well-differentiated adenosquamous carcinoma persisted in having a worse prognosis (58.3% ten-year survival rate), compared with adenocarcinoma (84.3% ten-year survival rate), which was explained by the propensity of adenosquamous carcinoma to deeply invade the myometrium.     

Abnormal distribution of E-cadherin and beta-catenin in different histologic types of cancer of the uterine cervix

OBJECTIVE: The goal of this study was to analyze the cellular distribution and possible alterations of beta-catenin and E-cadherin proteins in different histologic types of uterine cervical cancer and precursor lesions, compared to normal controls. METHODS: We performed an immunochemical staining analysis of the cellular distribution of beta-catenin and E-cadherin proteins in biopsy samples from 20 normal exocervical squamous epithelium, 43 premalignant lesions, and a large series of 126 invasive tumors of different histologic types that included 68 squamous carcinomas, 31 adenosquamous carcinomas, and 27 adenocarcinomas. Statistical significance was evaluated by the chi-square or Fisher's Exact test. RESULTS: We observed beta-catenin abnormally distributed in the cytoplasm of 62% of premalignant lesions and more than 70% of invasive cancers, statistically significant when compared with normal tissue (P < 0.05). Similarly, we found that E-cadherin exhibit a significant abnormal distribution in the cytoplasm of 58% of premalignant lesions (P < 0.05) and in more than 71% of squamous carcinoma and adenosquamous carcinoma when compared with normal tissue (P < 0.05). We found no differences in the distribution of E-cadherin between adenocarcinomas compared with control samples. Interestingly, we found that both, beta-catenin and E-cadherin, were absent in the membrane of nearly 40% premalignant lesions. Nuclear staining of beta-catenin was rarely seen in any cases, contrary to what has been reported for this and other neoplasias. CONCLUSION: Our findings indicate that cellular alterations of both beta-catenin and E-cadherin are frequent in tumors of the uterine cervix of different histologic types, and support a role for these proteins in cervical cancer development.     

Distribution of involucrin in normal and pathological human uterine cervix

A study was undertaken to determine the potential value of involucrin immunostaining, a protein synthesized by mature squamous epithelial cells, in distinguishing benign from neoplastic lesions in cervical pathology. A total of 146 cervical biopsies were analyzed using an indirect immunoperoxidase method and polyclonal antibody. A suprabasal homogeneous cytoplasmic staining pattern was consistently observed in normal squamous cervical epithelium. In contrast, 43.7% of cervical condylomas showed involucrin at all levels of the epithelium including the basal layer. Variable patterns were seen in cervical intraepithelial neoplasia (CIN), with 46% of full-thickness stainings, although no significant difference was obtained among the different grades of CIN lesions. Distribution of involucrin was correlated (P less than 0.05) with the degree of tumor differentiation in squamous cell carcinomas, being absent in 71.4% of poorly differentiated carcinomas and focally present in 75% of well-differentiated carcinoma. Lesions of endocervical origin, either benign or malignant, were entirely negative for involucrin. It is concluded that involucrin seems unable to establish a reliable differential diagnosis between benign and neoplastic conditions in cervical pathology, and should therefore be considered only a specific marker of squamous differentiation in both normal and pathological human uterine cervix.     

Evaluation of the Prognostic Significance of nm23/NDP Kinase Protein Expression in Cervical Carcinoma: An Immunohistochemical Study

The objective of this study was to evaluate the prognostic significance of immunohistochemical staining for nm23/nucleoside diphosphate (NDP) kinase in cervical carcinoma. A retrospective analysis of 176 patients with cervical carcinoma FIGO stage IB treated with radical hysterectomy and pelvic lymphadenectomy from 1987 to 1990 was conducted. Immunohistochemical staining using the polyclonal nm23-H1/NDP kinase A antibody was correlated to various histopathological and morphological characteristics (tumor size, histologic type, grade of differentiation, vessel invasion, invasion into parametria, and lymph node metastasis) and relapse-free survival. For controls, sections were obtained from 10 hysterectomy specimens with normal cervical epithelium. Staining for nm23/NDP kinase was observed in 90% of control cases and in 70.5% of cases of cervical carcinoma, more frequent in squamous and adenosquamous cell carcinoma than in adenocarcinoma and more frequent in poorly differentiated than in more highly differentiated tumors. There were no differences related to size of tumor or invasion into vessels or parametria or occurrence of lymph node metastasis. The relapse-free survival was lower for patients with squamous cell and adenosquamous tumors with positive immunostaining for nm23/NDP kinase than for those with negative tumors when evaluated in univariate analysis. In multivariate analysis with tumor size, vessel invasion, invasion into parametria, grade of differentiation, and lymph node metastasis included, this difference was no longer significant. In patients with adenocarcinoma no difference was found. In conclusion, we did not find immunostaining for nm23/NDP kinase to be a useful indicator for prognosis in cancer of the uterine cervix.     

Immunohistological demonstration of calcitonin in endometrial carcinomas with and without argyrophil cells

Sixteen endometrial carcinomas were studied by an immunoperoxidase method with anti-human calcitonin, 8 with and 8 without argyrophil cells. Calcitonin was demonstrated in 7 with and 2 without argyrophil cells. Calcitonin production was not associated with any specific histology of the endometrial carcinomas. Regardless of the existence of argyrophil cells, calcitonin was located in the apical portion or in the entire cytoplasm of the glandular tumor cells in the well differentiated adenocarcinomas. In a poorly differentiated adenocarcinoma and the adenosquamous carcinomas, it was found in many immature and squamous tumor cells as well as in some glandular tumor cells. The comparative studies between calcitonin and argyrophil granules in the endometrial carcinomas with argyrophil cells revealed that calcitonin was located in many tumor cells other than argyrophil cells. The extreme examples were 2 calcitonin-producing tumors without argyrophil cells. Calcitonin production of the endometrial carcinomas is a novel finding, but the biological significance of argyrophil cells remains to be further investigated.     

Endometrial extension of adenosquamous carcinoma of the uterine cervix

Endometrial in situ extension of cervical cancer is extremely uncommon. Previous reports only present the cases of squamous cell carcinoma or related category. This report presented adenosquamous carcinoma of the uterine cervix that showed a paradoxical extension of each component in a 72-year-old patient. Main tumor in the cervix was revealed to be adenosquamous carcinoma. The glandular component extended to the vagina, while the squamous component grew into the entire uterine cavity and replaced the glandular epithelium. We presented the first case of adenosquamous carcinoma of the uterine cervix with vaginal and endometrial extension. Furthermore, the endometrium was replaced with squamous component, while the vagina was invaded by glandular component. The observed paradoxical extension of the present case was extremely rare.     

Monoclonal antibodies for the histopathological diagnosis of cervical neoplasia

Five monoclonal antibodies (Ca1, HMFG 1 and 2, 8.30.3 and 77.1) were used to study the distribution of antibody binding sites in cervical tissue with a view to identifying a marker which would distinguish between benign and malignant cervical epithelium. Both benign tissue (mature and immature metaplastic squamous epithelium, congenital transformation zone and glandular epithelium) and neoplastic tissue (cervical intraepithelial neoplasia, 1, 2 and 3 and invasive squamous cell carcinoma) were stained by these antibodies. Although immature metaplastic epithelium stained strongly with all the antibodies, the intensity and distribution of staining in general did not distinguish between benign and neoplastic conditions. All five antibodies, raised against three different antigens, stained cervical tissue in a similar way and thus were unsuitable for use as specific tumour markers in equivocal cases. Further studies on other tumour markers are indicated in view of the potential value of this approach.     

Immunocytochemical localization of keratin in normal, dysplastic and neoplastic cervical epithelium

The PAP immunocytochemical technique utilizing specific keratin antibody was applied to paraffin sections from 36 cervical biopsies. Normal squamous epithelium and condylomas had similar patterns of keratin production with intense staining of intermediate and upper layers, while basal cells remained negative. Dysplasia, carcinoma in situ and infiltrating squamous carcinoma showed uneven distribution of keratin with the least amount seen in the areas with high mitotic rate and anaplasia. All large cell squamous carcinomas demonstrated presence of significant amounts of keratin. Squamous carcinomas of the small cell type were essentially keratin-free.     

Papillary serous carcinoma of the ovary with squamous differentiation

Two cases of serous carcinoma of the ovary with squamous differentiation are described. These neoplasms occurred in two women who were 63 and 46 years old and who presented with International Federation of Gynecology and Obstetrics (FIGO) stage IIB and III disease, respectively. The first tumor recurred following chemotherapy; radiotherapy was given, but the patient died of tumor at 4 years. The second neoplasm did not respond to chemotherapy and caused the patient's death at 2 years. In the first case, the squamous differentiation occurred as compact nests of ovoid to spindle cells intermixed with a papillary serous carcinoma component. The squamous component of the second case developed as single or small groups of enlarged, eosinophilic cells, often multinucleated, within nests of papillary serous carcinoma with typical psammoma bodies. In the latter case, intercellular bridges were identified by light microscopy. Both cases showed squamous features by immunohistochemistry, staining positively for 57-kd keratin, although in the first case but not the second there was also some weaker staining for 54-kd keratin, probably indicating incomplete squamous differentiation with retained glandular features. Electron microscopy in both cases revealed prominent cytoplasmic tonofilaments in the squamous component. These two cases reinforce the concept that squamous differentiation, although more frequent in some other types of common epithelial tumors, may occur in serous ovarian carcinomas as well.     

Mucoepidermoid carcinoma of uterine cervix stage IB. Long-term follow-up, histochemical and immunohistochemical study.

Twelve women with mucoepidermoid carcinoma of the cervix uteri were followed for 2-15 years after diagnosis. Three patients died within 14 months. All had lymph node metastases and/or vascular involvement and exhibited tumor invasion to a depth of 1.2-3.2 cm. Mucoepidermoid carcinoma is defined as a tumor with the appearance of squamous cell carcinoma without any glandular pattern and with demonstrable intracellular mucin. The mucin is best demonstrated by alcian blue and periodic acid-Schiff-diastase. In 265 cases of squamous cell carcinoma, stage IB, lymph node metastases were present in 14%. In the cases of mucoepidermoid carcinoma, the prevalence of nodal metastases was 33%. Because mucoepidermoid carcinomas appear to be more aggressive lesions than squamous cell carcinomas are, it may be advisable to stain all cervical squamous carcinomas for mucin if they demonstrate finely vacuolated cytoplasm and lack peripheral palisading. Immunohistochemical studies for carcinoembryonic antigen (CEA), keratin, and epithelial membrane antigen were positive in all tumors to varying degrees. The detection of CEA may be of additional help in establishing a diagnosis.     

Cervical carcinoma antigen: distribution in neoplastic lesions of the uterine cervix and comparison to other tumor markers

Four antibodies (anti-CCA, anti-CEA, Ca-l, and anti-EMA) were used to study the distribution of antibody-binding sites in normal endocervical mucosa, metaplastic squamous epithelium, squamous epithelium exhibiting varying grades of intraepithelial neoplasia, and invasive squamous cell carcinoma. Anti-CCA, a novel monoclonal antibody raised against an extract of squamous cell carcinoma of the cervix, recognizes dysplastic, neoplastic, and metaplastic cervical epithelial cells. While anti-CCA and Ca-l rarely stained normal glandular epithelium, 31.4 and 45.7% of the samples stained positively for CEA and EMA, respectively. There did not appear to be significant differences between anti-CCA and the other antibodies in the frequency with which neoplastic conditions were stained. Based upon these observations, it appears that none of the antibodies tested can be regarded as a specific tumor marker.     

Immunohistochemical localization of keratin and secretory component in carcinoma of the uterine cervix

Carcinoma of the uterine cervix is said to frequently show a combination of squamous epithelial and glandular epithelial characteristics. In the present study, immunohistochemical localization of keratin and secretory component (SC) was studied to clarify these characteristics of cancers of the cervix, and the following results were obtained. Demonstration of the localization of keratin and SC was useful in providing functional markers of the squamous and glandular epithelium of the cervix. In epidermoid carcinomas, the squamous epithelial character of the keratinizing carcinomas was strongest and decreased in the large cell non-keratinizing, followed by the small cell non-keratinizing carcinomas. The glandular character of these lesions decreased in the same order. Subclassification of CIS did not reveal any major changes with either kind of staining. So-called bipotential differentiation was found in 21% of the epidermoid, 53% of the adenocarcinomas and 13% of the CIS. In the clinical stages of epidermoid carcinomas, the stage I and II cases more frequently showed squamous characteristics than did the stage 0 cases.     

Immunohistochemical comparison of new monoclonal antibody 1C5 and carcinoembryonic antigen in the differential diagnosis of adenocarcinoma of the uterine cervix.

The reactivities of new monoclonal antibody 1C5 and anti-carcinoembryonic antigen (CEA) were determined immunohistochemically in 4 adenocarcinomas in situ, 20 invasive adenocarcinomas of various types, and 6 adenosquamous carcinomas of the uterine cervix, as well as in 10 endometrial adenocarcinomas and 10 normal cervices. Among the invasive adenocarcinomas, 90% were positively stained by 1C5 and 55% stained for CEA. Three of four in-situ adenocarcinomas were positively stained by 1C5 and two of four were positively stained by anti-CEA. All adenosquamous carcinomas were stained by 1C5 and four of six stained for CEA. Invasive adenocarcinomas always stained more intensely with 1C5 than did noninvasive lesions in the same specimen. Poorly differentiated adenocarcinomas were stained as strongly with 1C5 as were well-differentiated tumors, but CEA was less effective in identifying poorly differentiated lesions. 1C5 was also more useful than CEA was in distinguishing glandular from squamous neoplastic differentiation, and also appears to be useful in distinguishing endocervical from endometrial differentiation.     

Endocervical carcinoma: A study of 23 patients with clinical-pathological correlation

Twenty-three patients with endocervical carcinomas diagnosed at the Mount Sinai Medical Center during the last 5 years were reviewed (clinical history and histologic material). The tumors were endocervical-type adenocarcinomas, adenosquamous carcinoma, mucinous (colloid), papillary, and clear cell carcinomas. Eight associated carcinomas in situ were found. Younger patients tended to have more adenosquamous carcinomas, perhaps related to the frequency of squamous metaplasia in sexually active women. Twenty-two out of twenty-three patients were multiparous, many of them grand multiparas. In two patients the tumor followed parturition, and in two patients it was found in the cervical stump. No association with oral contraceptives was found. Multiparous women seem to be at increased risk for this malignancy. The early detection of endocervical carcinoma is based on the recognition of dysplasia and adenocarcinoma in situ in the biopsy material. This is of particular significance in the endocervical cancer because its location often prevents early detection.     

Association of syndecan-1 with tumor grade and histology in primary invasive cervical carcinoma

OBJECTIVES: The expression of syndecan-1, a cell surface heparan sulfate proteoglycan, is reduced during malignant transformation of squamous cells. Studies on squamous cell carcinoma of the head and neck have shown that syndecan-1-positive tumors are associated with longer overall and recurrence-free survival. The purpose of this study was to analyze syndecan-1 expression in invasive cervical carcinoma and to examine the association of syndecan-1 expression with prognostic factors and overall survival. METHODS: The study population consisted of 124 patients treated for primary invasive carcinoma of the uterine cervix at the Turku University Central Hospital during the years 1970-1988. The material consisted of 102 (82.3%) squamous cell carcinomas, 16 (12.9%) adenocarcinomas and 1 (0.8%) adenosquamous carcinoma, 1 (0.8%) small cell carcinoma, 1 (0. 8%) adenoid basal carcinoma, 1 (0.8%) carcinosarcoma, and 2 (1.6%) unclassified cervical carcinomas. Syndecan-1 expression was determined on paraffin-embedded tissue blocks using a human syndecan-1-specific monoclonal antibody B-B4 and immunohistochemistry. The expression of syndecan-1 was classified according to staining intensity as well as the percentage of positively stained tumor cells. RESULTS: Staining intensity was strong in 44 (36%) samples, while 24 (19%) specimens remained syndecan-1-negative. In 49 (40%) samples, the percentage of syndecan-1-positive cells was >/=90%. Syndecan-1 expression, as determined by >/=50% positively stained tumor cells, was associated with the grade of differentiation (P = 0.03) and squamous histology (P < 0.001), but was not associated with clinical stage (P = 0.16) or disease-free survival (P = 0.86). Age (P = 0.003) and clinical stage (P < 0.001) were significant prognostic factors, but syndecan-1 expression determined neither by percentage of positively stained tumor cells nor by staining intensity was associated with the outcome. CONCLUSIONS: In cervical carcinoma syndecan-1 is associated with histological differentiation grade and squamous histology, but does not predict clinical outcome.     

Regulation of apoptosis in squamous cell carcinoma of the vulva

OBJECTIVE: To analyze the expression of Bcl-2, Bax and ICH-1-L in squamous cell cancer of the vulva. STUDY DESIGN: Slides of 72 vulvar squamous cell carcinomas were stained immunohistologically for Bcl-2, Bax and ICH-1-L. They were analyzed for the percentage of positive tumor cells, staining intensity and pattern, and amount of Bcl-2-positive lymphocytes around the tumor. Results were analyzed for correlations with clinical and histologic characteristics. Disease-free and overall survival were evaluated by Kaplan-Meier curves with the log-rank test. RESULTS: Strong expression of Bcl-2 was present in 15% of tumors. Carcinomas with high Bcl-2 expression more frequently had lymph node metastasis (P = .03), without significant differences in other clinical or histologic parameters, disease-free and overall survival. Strong Bax expression was observed in 57%, without prognostic significance. Carcinomas showed high ICH-1-L expression in 35%. These tumors seemed to have longer disease-free survival, while overall survival was significantly longer (P = .02). A strong Bcl-2-positive inflammatory infiltrate was highly predictive of lymph node metastasis (P = .02) and disease-free survival (P = .03). CONCLUSION: In squamous cell carcinoma of the vulva, inhibition of apoptosis is associated with a more-aggressive phenotype, and a Bcl-2-positive inflammatory infiltrate is predictive of prognosis. A study with more patients should confirm the importance of apoptosis in vulvar carcinoma.     

Human papillomavirus deoxyribonucleic acid in adenocarcinoma and adenosquamous carcinoma of the uterine cervix

This report describes the detection of human papillomavirus type 16 or 18 deoxyribonucleic acid (DNA) in nine of 15 invasive tumors of the cervix, including three squamous carcinomas, four adenosquamous carcinomas, one glassy cell carcinoma, and one adenocarcinoma. The viral DNA was identified by Southern blotting and DNA hybridization. Human papillomaviruses may play an etiologic role in the development of at least some adenocarcinomas and adenosquamous carcinomas as well as most squamous tumors of the cervix.     

Concomitant endometrial adenoacanthoma and bilateral (inguinal) lymph node endometriotic adenoacanthomas or nodal metastases of the endometrial adenoacanthoma? A case report with a literature survey of the histogenetic aspects of endometriotic foci in pelvic lymph nodes

This case report depicts an interesting association of well-differentiated adenocarcinoma of the endometrium with benign squamous metaplastic foci confined to the fundus uteri, superficially invading the myometrium and with concomitant bilateral pelvic lymph node endometriotic adenoacanthomas. Right inguinal lymphadenopathy was detected during the first hospitalization. A lymph node biopsy from the right groin, carried out at Nahariyya, revealed apparent metastatic adenoacanthoma regarded most probably as endometrial in origin. Fractionated curettage later showed a stage Ia G1 adenocarcinoma of the endometrium with benign squamous metaplastic elements (adenoacanthoma). At Rambam Hospital, Haifa, left groin node enlargement was noted as well. The Gynecologic Oncology Unit confirmed the previous histologic findings. At exploratory laparotomy total abdominal hysterectomy and bilateral salpingo-oophorectomy, paraaortic, and bilateral pelvic lymph node sampling as well as appendectomy were performed. Bilateral groin node dissection was then carried out. Out of 37 nodes examined tumor was found in only one node, namely that of the left groin. The paper includes an extensive survey of the literature on the subject of the histopathogenesis of endometriotic foci in pelvic lymph nodes and the discussion is designed to elucidate the diagnostic problem involved in this case report.