An Immunohistochemical Algorithm for Ovarian Carcinoma Typing.

There are 5 major histotypes of ovarian carcinomas. Diagnostic typing criteria have evolved over time, and past cohorts may be misclassified by current standards. Our objective was to reclassify the recently assembled Canadian Ovarian Experimental Unified Resource and the Alberta Ovarian Tumor Type cohorts using immunohistochemical (IHC) biomarkers and to develop an IHC algorithm for ovarian carcinoma histotyping. A total of 1626 ovarian carcinoma samples from the Canadian Ovarian Experimental Unified Resource and the Alberta Ovarian Tumor Type were subjected to a reclassification by comparing the original with the predicted histotype. Histotype prediction was derived from a nominal logistic regression modeling using a previously reclassified cohort (N=784) with the binary input of 8 IHC markers. Cases with discordant original or predicted histotypes were subjected to arbitration. After reclassification, 1762 cases from all cohorts were subjected to prediction models (χ Automatic Interaction Detection, recursive partitioning, and nominal logistic regression) with a variable IHC marker input. The histologic type was confirmed in 1521/1626 (93.5%) cases of the Canadian Ovarian Experimental Unified Resource and the Alberta Ovarian Tumor Type cohorts. The highest misclassification occurred in the endometrioid type, where most of the changes involved reclassification from endometrioid to high-grade serous carcinoma, which was additionally supported by mutational data and outcome. Using the reclassified histotype as the endpoint, a 4-marker prediction model correctly classified 88%, a 6-marker 91%, and an 8-marker 93% of the 1762 cases. This study provides statistically validated, inexpensive IHC algorithms, which have versatile applications in research, clinical practice, and clinical trials.

HNF-1ß in ovarian carcinomas with serous and clear cell change.

Many ovarian tumors, including high-grade serous carcinoma (HGSC), show clear cell change. Accurate diagnosis is important, however, as ovarian clear cell carcinoma (OCCC) is known to be less responsive to traditional types of ovarian cancer chemotherapies. In a previous study, the clinical, morphologic, and immunohistochemical features of 32 ovarian carcinomas, which had been previously diagnosed as pure OCCC (n=11), pure HGSC (n=11), and mixed serous and clear cell (MSC) (n=10), were analyzed. The immunoreactivities of WT1, ER, and p53, as well as the mitotic indices and stages of presentation of the MSC, were similar to those of HGSC. It was consequently concluded that MSC represented HGSC with clear cell change. Hepatocyte nuclear factor-1ß (HNF-1ß) is a relatively new immunohistochemical marker that has been shown to be rather sensitive and specific for OCCC. We thus sought to evaluate this marker in this specific group of tumors. One block each of pure HGSC and pure OCCC were stained with HNF-1ß. In the cases of MSC, 2 blocks were stained when the serous and clear cell components were not present on the same slide. None (0/11) of the pure HGSC showed immunoreactivity for HNF-1ß, whereas all (11/11) of the pure OCCC were positive. In the cases of MSC, both the serous and clear cell components were negative for HNF-1ß. HNF-1ß seems to be a sensitive and specific marker for OCCC and is not expressed in HGSC with clear cell change. The pattern of immunoreactivity of HNF-1ß in tumors with both serous and clear cell change supports the conclusion that MSC are HGSC with clear cells.

Reproducibility of histological cell type in high-grade endometrial carcinoma.

Subclassification of endometrial carcinoma according to histological type shows variable interobserver agreement. The aim of this study was to assess specifically the interobserver agreement of histological type in high-grade endometrial carcinomas, recorded by gynecological pathologists from five academic centers across Canada. In a secondary aim, the agreement of consensus diagnosis with immunohistochemical marker combinations was assessed including six routine (TP53, CDKN2A (p16), ER, PGR, Ki67, and VIM) and six experimental immunohistochemical markers (PTEN, ARID1A, CTNNB1, IGF2BP3, HNF1B, and TFF3). The paired interobserver agreement ranged from κ 0.50 to 0.63 (median 0.58) and the intraobserver agreement from κ 0.49 to 0.67 (median 0.61). Consensus about histological type based on morphological assessment was reached in 72% of high-grade endometrial carcinomas. A seven-marker immunohistochemical panel differentiated FIGO grade 3 endometrioid from serous carcinoma with a 100% concordance rate compared with the consensus diagnosis. More practically, a three-marker panel including TP53, ER, and CDKN2A (p16) can aid in the differential diagnosis of FIGO grade 3 endometrioid from endometrial serous carcinoma. Our study demonstrates that the inter- and intraobserver reproducibility of histological type based on morphology alone are mostly moderate. Ancillary techniques such as immunohistochemical marker panels are likely needed to improve diagnostic reproducibility of histological types within high-grade endometrial carcinomas.

Diagnosis of ovarian carcinoma cell type is highly reproducible: a transcanadian study.

Reproducible diagnosis of ovarian carcinoma cell types is critical for cell type-specific treatment. The purpose of this study was to test the reproducibility of cell type diagnosis across Canada. Analysis of the interobserver reproducibility of histologic tumor type was performed among 6 pathologists after brief training in the use of modified World Health Organization criteria to classify ovarian carcinomas into 1 of 6 categories: high-grade serous, endometrioid, clear cell, mucinous, low-grade serous, and other. These 6 pathologists independently reviewed a test set of 40 ovarian carcinomas. A validation set of 88 consecutive ovarian carcinomas drawn from 5 centers was subject to local review by 1 of the 6 study pathologists, and central review by a single observer. Interobserver agreement was assessed through calculation of concordance and kappa values for pair-wise comparison. For the test set, the paired concordance between pathologists in cell type diagnosis ranged from 85.0% to 97.5% (average 92.3%), and the kappa values were 0.80 to 0.97 (average 0.89). Inclusion of immunostaining results did not significantly improve reproducibility (P=0.69). For the validation set, the concordance between original diagnosis and local review was 84% and between local review and central review was 94%. The kappa values were 0.73 and 0.89, respectively. With a brief training exercise and the use of defined criteria for ovarian carcinoma subtyping, there is excellent interobserver reproducibility in diagnosis of cell type. This has implications for clinical trials of subtype-specific ovarian carcinoma treatments.

IGF2BP3 (IMP3) expression is a marker of unfavorable prognosis in ovarian carcinoma of clear cell subtype.

Clear cell carcinoma is an uncommon subtype of ovarian carcinoma, accounting for 10% of cases. Clear cell carcinoma typically presents with stage I or II disease, and in this setting prognostic markers could aid in management decisions, in particular the decision to treat with adjuvant chemotherapy. We tested whether expression of insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3, also known as IMP3) can serve as a new biomarker to predict outcome for patients with clear cell carcinoma and other subtypes of ovarian carcinoma. The expression of IGF2BP3 was evaluated by immunohistochemistry in 475 ovarian carcinomas of different subtypes and correlated with disease-specific survival. IGF2BP3 antibody specificity was validated by correlation of IGF2BP3 protein with mRNA expression level in a series of 35 ovarian carcinomas (r=0.849, P<0.0001). IGF2BP3 protein expression was an independent marker of reduced disease-specific survival (risk ratio 2.9, 95% confidence interval 1.4-5.8) in the clear cell subtype (N=128), but not in high-grade serous (N=198) or endometrioid (N=121) carcinomas. The prognostic significance of IGF2BP3 expression for reduced disease-specific survival (risk ratio 2.6, 95% confidence interval 1.3-5.0) was confirmed in an independent series of cases (N=150) from three different centers in North America. We conclude that IGF2BP3 is the first biomarker of prognostic significance in ovarian clear cell carcinoma that has been validated in an independent case series.

A limited panel of immunomarkers can reliably distinguish between clear cell and high-grade serous carcinoma of the ovary.

The distinction of ovarian clear cell carcinomas (CCCs) from high-grade serous carcinomas (HG-SCs) is sometimes a diagnostic challenge. With the recognition that CCCs respond poorly to conventional chemotherapy there are efforts to initiate clinical trials for CCC, making accurate diagnosis critical. The purpose of this study was to test and validate a set of antibodies that could aid in the diagnosis of CCC, using a series of cases from different centers in North America. Using a test set of 133 CCCs, we identified the following markers: Cyclin E, estrogen receptor, hepatocyte nuclear factor (HNF)-1beta, Ki-67, p21, p53, and Wilms tumor (WT)1 that show significant discrimination from 200 HG-SCs. For validation, these markers were characterized on an independent set of 104 CCCs from 3 other centers. There were no significant differences in expression of these 7 markers between the independent test and validation sets of CCC. Combining all CCC cases (N=237), HNF-1beta showed the highest sensitivity (82.5%) and specificity (95.2%) for CCC, and WT1 for HG-SC (sensitivity: 79.9%, specificity: 97.4%). A diagnostic panel consisting of WT1, ER, and HNF-1beta demonstrated nearly identical performance as a panel using all 7 markers in distinguishing CCCs from HG-SCs, correctly classifying 84% of cases. Three percent of cases were misclassified and 13% carried an uninformative triple negative immunophenotype. CCCs show a distinct, reproducible immunophenotype, compared with HG-SCs, and a panel of 3 immunomarkers can serve as a diagnostic aid in problematic cases.

Mixed ovarian epithelial carcinomas with clear cell and serous components are variants of high-grade serous carcinoma: an interobserver correlative and immunohistochemical study of 32 cases.

There are conflicting data about chemoresistance and prognosis in ovarian clear cell carcinoma (CCC). This could be due to significant interobserver variation in the diagnosis of CCC and other ovarian surface epithelial tumors containing clear cells. Thirty-two cases previously diagnosed as CCC, high-grade ovarian serous carcinoma (SC), and mixed surface epithelial carcinoma (SEC) with clear cell and serous components were reviewed by 4 gynecologic pathologists blinded to the original diagnoses. Interobserver reproducibility was evaluated. Each case was also assessed using immunohistochemical markers Wilm tumor 1, estrogen receptor, and p53. The interobserver reproducibility was greatest for pure CCC (kappa of 0.82), and lowest for the mixed SEC (kappa of 0.32). Moderate agreement was seen in the pure SC category (kappa of 0.59). All pure SC and most mixed SEC presented as stage III or IV diseases. Most pure CCC presented as stage I or II diseases. Immunoreactivities of the mixed SECs were similar to those of pure SC, but significantly different from those of pure CCC for Wilm tumor 1 (P=0.0011 for both components), estrogen receptor (P=0.0003 for clear cell component, P=0.0001 for serous component), and p53 (P=0.0062 for both components). The serous and clear cell components of mixed SEC showed higher mitotic rates than pure CCC (P=0.004 and P=0.023, respectively), but the mitotic rate of pure SC was similar to the mixed SEC. We conclude that (1) pure CCC is reproducibly diagnosed. (2) The diagnosis of mixed ovarian SEC with clear cell component is not reproducible. (3) Mixed SEC with clear cell and serous components show similar stage, mitotic activities, and immunoreactivities to those of pure SC, and likely represent SC with clear cell changes.