Inclusion of MUC1 (Ma695) in a panel of immunohistochemical markers is useful for distinguishing between endocervical and endometrial mucinous adenocarcinoma.

BACKGROUND: Distinguishing endocervical adenocarcinoma (ECA) from endometrial mucinous adenocarcinoma (EMMA) is clinically significant in view of the differences in their management and prognosis. In this study, we used a panel of tumor markers to determine their ability to distinguish between primary endocervical adenocarcinoma and primary endometrial mucinous adenocarcinoma. METHODS: Immunohistochemistry using monoclonal antibodies to MUC1 (Ma695), p16, estrogen receptor (ER), progesterone receptor (PR), and vimentin, was performed to examine 32 cases, including 18 EMMAs and 14 ECAs. For MUC1, cases were scored based on the percentage of staining pattern, apical, apical and cytoplasmic (A/C), or negative. For p16, cases were scored based on the percentage of cells stained. For the rest of the antibodies, semiquantitative scoring system was carried out. RESULTS: For MUC1, majority of EMMA (14 of 18 cases, 78%) showed A/C staining, whereas only few ECA (2 of 14, 14%) were positive. The difference of MUC1 expression in the two groups of malignancy was statistically significant (p < 0.001). Staining for p16 was positive in 10 of 14 (71%) ECA and 4 of 18 (22%) EMMA. Estrogen receptor was positive in 3 of 14 (21%) ECA and 17 of 18 (94%) EMMA. Progesterone receptor was positive in 3 of 14 (21%) ECA and 16 of 18 (89%) EMMA. Vimentin was positive in 1 of 14 (7%) ECA, and 9 of 18 (50%) EMA, with median and range of 0 (0-6), and 1.5 (0-9) respectively. CONCLUSION: A panel of immunohistochemical markers including MUC1, p16, ER, PR, and vimentin is recommended, when there is morphological and clinical doubt as to the primary site of endocervical or endometrial origin.

Expression of Wilms tumor gene (WT1) in epithelial ovarian cancer.

OBJECTIVES: The identification of proteins that are selectively expressed in cancer and with potential to elicit an immune response is the first step towards antigen-specific immunotherapy. The Wilms tumor gene product (WT1) is inherently immunogenic and is now thought to be oncogenic. The aim of this study was to determine the expression of WT1 in epithelial ovarian cancer (EOC) and correlate with clinico-pathologic characteristics. METHODS: WT1 expression was examined using immunohistochemistry applied on a tissue microarray of normal tissues and a panel of 100 EOC tissues. The distribution of WT1 expression and clinico-pathologic variables were analyzed. Survival probabilities were estimated by Kaplan-Meier method, and statistical significance was determined by the log-rank test. RESULTS: WT1 expression was observed in 78/100 of specimens. The predominant expression pattern was homogenous, occurring in 66/100 (66%) of WT1-positive specimens, while 12/100 (12%) demonstrated heterogeneous staining. In normal tissues, WT1 expression was noted in kidneys, splenic capsule, Sertoli cells of the testis, and granulosa cells of the ovary. The median follow-up of the patient population was 30 months. Patients with WT1-positive tumors tended to have a higher grade (P = 0.006) and stage (P = 0.002) of tumor. However, there were no significant differences in the distribution of patients with WT1-positive tumors in relation to disease-free and overall survival. CONCLUSIONS: Our data demonstrate that WT1 is expressed at high frequency in patients with EOC. Since WT1 demonstrates tissue-restricted expression and is inherently immunogenic, it could represent an attractive target for antigen-specific immunotherapy in EOC.

Detection of epithelial ovarian cancer using 1H-NMR-based metabonomics.

Currently available serum biomarkers are insufficiently reliable to distinguish patients with epithelial ovarian cancer (EOC) from healthy individuals. Metabonomics, the study of metabolic processes in biologic systems, is based on the use of (1)H-NMR spectroscopy and multivariate statistics for biochemical data generation and interpretation and may provide a characteristic fingerprint in disease. In an effort to examine the utility of the metabonomic approach for discriminating sera from women with EOC from healthy controls, we performed (1)H-NMR spectroscopic analysis on preoperative serum specimens obtained from 38 patients with EOC, 12 patients with benign ovarian cysts and 53 healthy women. After data reduction, we applied both unsupervised Principal Component Analysis (PCA) and supervised Soft Independent Modeling of Class Analogy (SIMCA) for pattern recognition. The sensitivity and specificity tradeoffs were summarized for each variable using the area under the receiver-operating characteristic (ROC) curve. In addition, we analyzed the regions of NMR spectra that most strongly influence separation of sera of EOC patients from healthy controls. PCA analysis allowed correct separation of all serum specimens from 38 patients with EOC (100%) from all of the 21 premenopausal normal samples (100%) and from all the sera from patients with benign ovarian disease (100%). In addition, it was possible to correctly separate 37 of 38 (97.4%) cancer specimens from 31 of 32 (97%) postmenopausal control sera. SIMCA analysis using the Cooman's plot demonstrated that sera classes from patients with EOC, benign ovarian cysts and the postmenopausal healthy controls did not share multivariate space, providing validation for the class separation. ROC analysis indicated that the sera from patients with and without disease could be identified with 100% sensitivity and specificity at the (1)H-NMR regions 2.77 parts per million (ppm) and 2.04 ppm from the origin (AUC of ROC curve = 1.0). In addition, the regression coefficients most influential for the EOC samples compared to postmenopausal controls lie around delta3.7 ppm (due mainly to sugar hydrogens). Other loadings most influential for the EOC samples lie around delta2.25 ppm and delta1.18 ppm. These findings indicate that (1)H-NMR metabonomic analysis of serum achieves complete separation of EOC patients from healthy controls. The metabonomic approach deserves further evaluation as a potential novel strategy for the early detection of epithelial ovarian cancer.

Somatic D-loop mitochondrial DNA mutations are frequent in uterine serous carcinoma.

The mitochondria plays a role in apoptosis. Its genome is also more susceptible to mutations because of high levels of reactive oxygen species and limited repair mechanisms. The D-loop of mitochondrial DNA (mtDNA) contains essential transcription and replication elements, and mutations in this region might alter the rate of DNA replication. We examined genetic alterations in the D-loop region of mtDNA in uterine serous carcinoma (USC) samples and their paired normal adjacent endometrium. DNA was extracted after laser-capture microdissection of paraffin-embedded tissues from eight patients with USC. The entire D-loop genome was amplified using nine pairs of overlapping primers. Denatured polymerase chain reaction (PCR) products were subjected to single-strand conformation polymorphism (SSCP) analysis. Somatic mtDNA alterations were detected in five tumours (63%). Our study indicates that mtDNA D-loop sequence alterations occur at a high frequency in USC suggesting that mtDNA mutations may play a role in the development of USC.

NY-ESO-1 and LAGE-1 cancer-testis antigens are potential targets for immunotherapy in epithelial ovarian cancer.

Cancer-testis (CT) antigens are expressed in a variety of cancers, but not in normal adult tissues, except for germ cells of the testis, and hence appear to be ideal targets for immunotherapy. In an effort to examine the potential of NY-ESO-1 and LAGE-1 CT antigens for immunotherapy in epithelial ovarian cancer (EOC), we examined the expression of these antigens by reverse transcription-PCR (RT-PCR) and immunohistochemistry (IHC) in a large panel of EOC tissues and cell lines. Sera from a subgroup of the patients were tested for NY-ESO-1/LAGE-1 antibody by ELISA. The data indicated that four ovarian cancer cell lines were positive for one or both CT antigens. Expression of NY-ESO-1 in EOC was demonstrated by RT-PCR and/or IHC in 82 of 190 (43%) specimens. NY-ESO-1 expression by IHC ranged from homogeneous to heterogeneous pattern. LAGE-1 mRNA expression was present in 22 of 107 (21%) tumor tissues. Overall, the expression of either NY-ESO-1 or LAGE-1 mRNA was present in 42 of 107 (40%) EOC specimens and coexpression of both antigens was demonstrated in 11% of specimens. Antibody to NY-ESO-1/LAGE-1 was present in 11 of 37 (30%) patients whose tumors expressed either NY-ESO-1 or LAGE-1. Detectable antibodies were present for up to 3 years after initial diagnosis. Although there was no statistically significant relation between expression of NY-ESO-1/LAGE-1 antigen and survival, the data showed aberrant expression of NY-ESO-1 and LAGE-1 by IHC/RT-PCR in a significant proportion of EOC patients. These findings indicate that NY-ESO-1 and LAGE-1 are attractive targets for antigen-specific immunotherapy in EOC.