PI3K Pathway Effectors pAKT and FOXO1 as Novel Markers of Endometrioid Intraepithelial Neoplasia The diagnosis of endometrioid intraepithelial neoplasia (EIN) is challenging owing to limited sampling, hormonal status, and other confounding histologic variables. Markers such as PTEN or PAX2 can delineate EIN in some cases, but are not wholly reliable. Clearly, new markers of EIN are needed. We explored several potential markers of EIN based rationally on molecular pathways most frequently misregulated in endometrial cancer: the 3-phosphoinositide kinase (PI3K)/AKT, β-catenin, and mismatch repair pathways. We studied PTEN, PAX2, β-catenin, and MLH1, in conjunction with 2 new markers—FOXO1 and phosphorylated AKT (pAKT)—not previously investigated in EIN. Benign (n=14) and EIN (n=35) endometria were analyzed by immunohistochemistry. Staining patterns were interpreted, tabulated, and scored by “clonal distinctiveness” in neoplastic lesions; that is, pattern alterations relative to normal glands. In normal endometria, FOXO1 was cytoplasmic in proliferative phase, but nuclear in secretory phase, showing that PI3K/FOXO1 participates in endometrial cycling and that FOXO1 is a readout of PI3K status. pAKT expression was low across normal endometria. FOXO1 or pAKT expression was altered in the majority of EINs (27/35, 77%), with FOXO1 and pAKT being co-altered only in some (20/35, 57%). β-catenin or MLH1 also exhibited clonal distinctiveness in EINs, showing that these are also useful markers in some cases. This is the first study to demonstrate the potential of pAKT and FOXO1 as biomarkers in the histopathologic evaluation of EIN. However, variability in expression poses challenges in interpretation.

Audit of Endometrial Cancer Pathology for a Regional Gynecological Oncology Multidisciplinary Meeting Endometrial cancer is a common disease, and in England all cancer cases are discussed at a central multidisciplinary meeting (MDT) with pathology review. We reviewed cases discussed/reviewed at a regional Gynecology MDT comparing (i) original referral histology with review histology and (ii) final review histology with the final hysterectomy histology. Cases identified as potentially eligible for the study (n=884) were found over a 4-yr period. This was reduced to 630 due to data and other issues for the primary biopsy review, and to 488 for both biopsy and hysterectomy sample. Cases were classed by agreement by grade/type and compared by clinical management (low grade vs. high grade). Of the original biopsies, central review agreed exactly with 67% and disagreed with 33%. A total of 11.6% of low-grade cancers were upgraded to high grade on review, and 6.1% of high-grade cancers were downgraded. For the biopsy/hysterectomy comparison, this was 72.5% agreement and 27.5% disagreement, with 3.5% upgraded to high from low grade, and 7.5% downgraded from high to low grade. The main areas of significant change was the identification of high-grade serous carcinoma from low-grade endometrial cancers, as well some other high grade types (clear cell and carcinosarcoma) and the confident diagnosis of cancer as opposed to an atypical hyperplasia. Central pathology review for MDT discussion does highlight significant areas of pathologic disagreement that would affect clinical management. The audit highlights that a significant disagreement rate in reporting such material between pathologists may be inescapable, but can be reduced by review.

Histologic Features of Hysterectomy Specimens From Female-Male Transgender Individuals Histologic changes in the female genital tract after prolonged androgen stimulation have been described in the past. However, these changes have not been systematically addressed in hysterectomy specimens from subjects undergoing surgical gender-reassignment, typically after treatment with exogenous androgens. The current study aims to provide practicing pathologists with a list of expected histologic features in hysterectomy specimens from female-male transgender individuals. Twenty-seven hysterectomy with bilateral salpingo-oophorectomy specimens were identified from our Laboratory Information System. Slides were retrieved and reviewed for features associated with androgen exposure. Clinical information for the 27 subjects (20–46 yr old, mean=29 yr) was obtained from the electronic medical records. Twenty-four subjects had received androgen 19 mo to 24 yr preoperatively. Focal decidua-like endometrial stromal change with glandular paucity was present in 16/27 (59%) uteri associated with predominantly inactive endometrial glands. Ectocervical or transformation zone transitional cell metaplasia was present in 17/27 (63%) subjects. Bilateral cystic follicles were present in all 23 subjects who underwent bilateral salpingo-oophorectomy and had preoperative androgen exposure. In these ovaries, follicular density appeared higher than that expected for age with counts ranging from 1.5 to 32.5 follicles/mm2 (average=10.7 follicles/mm2). Predominantly inactive, sparse endometrial glands with focal decidua-like stromal change, cervical transitional cell metaplasia, bilateral cystic follicles and higher follicular density are observed in the majority of specimens from female-male transgender individuals. These histologic changes correlate with prolonged preoperative androgen administration. The significance of these findings relies on recognizing the spectrum of androgen-related histologic alterations and not confusing transitional cell metaplasia with cervical dysplasia.

YWHAE-NUTM2A/B Translocated High-grade Endometrial Stromal Sarcoma Commonly Expresses CD56 and CD99 YWHAE-NUTM2A/B translocated high-grade endometrial stromal sarcoma is an uncommon and clinically aggressive uterine mesenchymal neoplasm which is sometimes associated with a low-grade fibromyxoid component. Although the diagnosis is confirmed using molecular techniques, it is possible to make the diagnosis based on the characteristic morphology and the immunophenotype; the latter typically includes diffuse positive staining of the high-grade component with cyclin D1 and CD117 and negative staining with CD10 and hormone receptors. Especially in a small biopsy specimen, in cases without a low-grade component or when presenting with extrauterine disease, the differential diagnosis may be wide and potentially include “small round cell tumors” such as a high-grade neuroendocrine carcinoma and a tumor in the Ewing family. In this study, we report consistent CD56 and CD99 immunoreactivity in YWHAE-NUTM2A/B translocated high-grade endometrial stromal sarcoma. We stained 20 of these neoplasms (10 molecularly confirmed and 10 where diagnosis was based on the morphology and immunophenotype) with CD56 and CD99; in one case, CD56 staining was not performed. Nineteen of 19 (100%) and 17 of 20 (85%) were positive with CD56 and CD99, respectively. Staining was usually diffuse with membranous and cytoplasmic immunoreactivity. Pathologists should be aware that YWHAE-NUTM2A/B translocated high-grade endometrial stromal sarcoma is commonly positive with CD56 and CD99; as well as potentially assisting in diagnosis, positivity with these markers may also result in consideration of other neoplasms in the differential diagnosis.

Mismatch Repair Protein Expression in Endometrioid Intraepithelial Neoplasia/Atypical Hyperplasia: Should We Screen for Lynch Syndrome in Precancerous Lesions? Screening for Lynch syndrome (LS) is routinely performed in patients with endometrial carcinoma. Currently, no screening recommendations exist for LS in precancerous lesions. The study goal was to determine the incidence of abnormal protein expression in endometrioid intraepithelial neoplasia/atypical hyperplasia (EIN/AH). We analyzed mismatch repair (MMR) protein expression by immunohistochemistry in EIN/AH concurrent with MMR-deficient endometrial carcinomas, and in endometrial biopsy/curettage specimens with EIN/AH from an unselected group of patients. Of 63 patients with MMR-deficient endometrial carcinoma, 34 demonstrated loss of MLH1/PMS2 expression; 1 showed loss of PMS2 alone; 12 showed loss of MSH2/MSH6, and 15 had loss of MSH6 alone. Genetic testing identified deleterious mutations in 14 cases (LS). 15 tumors demonstrated MLH1 promoter hypermethylation. Abnormal MMR expression in EIN/AH and adjacent carcinoma was concordant in 100% of LS cases and 71% of MLH1 promoter hypermethylation cases. Of 118 patients from the unselected group with EIN/AH, 4 (3%) cases demonstrated absent expression of one or more MMR proteins. Of these, 2 patients were later confirmed to have deleterious mutations in subsequent specimens with endometrial carcinoma. The prevalence of abnormal MMR expression in EIN/AH adjacent to carcinoma and in the unselected group of patients with EIN/AH is similar to the reported prevalence of LS in endometrial carcinoma. Identifying patients at high risk for LS through abnormal MMR expression in EIN/AH provides the benefit of early surveillance, treatment and timely diagnosis for the patient and affected family members.

Targeted Genomic Profiling of Female Adnexal Tumors of Probable Wolffian Origin (FATWO) Female adnexal tumor of probable Wolffian origin (FATWO) is a rare gynecologic neoplasm of low-malignant potential presumed to be derived from mesonephric remnants in the upper female genital tract. Similarly, mesonephric remnants in the lower female genital tract are thought to be the origin for mesonephric carcinoma. Although the molecular alterations in mesonephric carcinoma have been recently reported, the pathogenesis of and molecular alterations in FATWO are not well understood. The aims of this study were to examine the molecular alterations in FATWO and to establish whether these neoplasms are molecularly similar to mesonephric carcinoma. Eight FATWOs underwent massively parallel sequencing to detect single nucleotide variations, copy number variations, and structural variants by surveying exonic DNA sequences of 300 cancer genes and 113 introns across 35 genes. Good quality DNA was isolated from 7 of 8 cases. Novel KMT2D variants (1 frameshift, 3 missense) were identified in 4 of 7 cases (57%), but were variants of uncertain biologic significance. STK11 mutations (both frameshift) were identified in 2 of 7 cases (29%); one of these was in a patient with a known history of Peutz-Jeghers syndrome. A mutation in the chromatin remodeling gene ARID1B was identified in 1 of 7 cases (14%). No cases harbored KRAS, NRAS, TP53, PIK3CA, PTEN, or DICER1 mutations. There were relatively low numbers of copy number variations, and no recurrent copy number variations were identified. One case demonstrated moderate copy gain of CCND1. No structural variants were identified. In summary, FATWO is characterized molecularly by the absence of KRAS/NRAS mutations (characteristic of mesonephric carcinoma), absence of DICER1 mutations (characteristic of Sertoli-Leydig cell tumor) and frequent KMT2D mutations of unknown biologic significance. FATWOs exhibit a limited number of molecular aberrations that are significantly different from those reported in tumors in the differential diagnosis, and our results question the relationship of mesonephric carcinoma with FATWO. Disease-defining molecular alterations for FATWO have yet to be discovered.

Tubo-Ovarian Transitional Cell Carcinoma and High-grade Serous Carcinoma Show Subtly Different Immunohistochemistry Profiles Tubo-ovarian transitional cell carcinoma (TCC) is grouped with high-grade serous carcinoma (HGSC) in the current World Health Organization classification. TCC is associated with BRCA mutations and a better prognosis compared with HGSC. Previous papers examining the immunohistochemical features of TCC have studied limited numbers of samples. No marker reflecting the biological difference between TCC and HGSC is known. We collected a large cohort of TCC to determine whether TCC and HGSC could be distinguished by immunohistochemistry. A tissue microarray was built from 89 TCC and a control cohort of 232 conventional HGSC. Immunohistochemistry was performed, scored, and statistically analyzed for routine markers of HGSC and urothelial tumors: PAX8, WT1, p53, p16, ER, p63, and GATA3. Using scoring cutoffs commonly employed in clinical practice, the immunohistochemical profile of TCC was indistinguishable from HGSC for all markers. However, more detailed scoring criteria revealed statistically significant differences between the 2 groups of tumors with respect to ER, PAX8, and WT1. HGSC showed more diffuse and intense staining for PAX8 (P=0.004 and 0.001, respectively) and WT1 (P=0.002 and 0.002, respectively); conversely, TCC showed more intense staining for ER (P=0.007). TCC and HGSC therefore show subtle differences in their immunohistochemical profiles which might reflect underlying (epi)genetic differences. Further studies using proteomic analysis will focus on the identification of differentially expressed proteins that might serve as markers of TCC-like differentiation, which could help explain biologic differences between TCC and HGSC and might identify other cases of HGSC with a better prognosis.

Multiple Luteinized Follicle Cysts of the Ovary in a Patient With a Pituitary Adenoma: Report of a Case and Review of the Literature The case of a 36-yr-old woman with a pituitary adenoma who was found to have bilateral ovarian masses is reported. The right ovary was removed, measured 15 cm in maximum dimension, and contained multiple cysts which on microscopic examination had the typical morphology of follicle cysts. The left ovary was grossly similar intraoperatively. Subsequent excision of the pituitary adenoma was followed ∼3 mo later by a return to normal size of the left ovary. The case represents an example of multiple luteinized follicle cysts, analogous to the phenomenon seen occasionally in pregnancy, but with a different clinical background. Periodic documentation of this phenomenon is present in the literature, predominantly the clinical literature with limited pathologic documentation of the nature of the process in many reports. As pertains to the evaluation of follicle cysts encountered during pregnancy the differential diagnosis is with a cystic granulosa cell tumor of either adult or juvenile types, more likely the latter. The cyst lining is identical to that of standard follicle cysts and contrasts with the immature mitotically active nuclei seen in a juvenile granulosa cell tumor. That neoplasm also usually shows follicular differentiation typically absent in follicle cysts. Pathologists should be aware of the rare occurrence of luteinized follicle cysts in patients with a pituitary adenoma to enable correct intraoperative and standard pathologic evaluation.

Ovarian Endometrioid Carcinoma Misdiagnosed as Mucinous Carcinoma: An Underrecognized Problem Primary mucinous carcinoma of the ovary is uncommon, and while numerous studies have focused on improving our ability to distinguish these tumors from gastrointestinal metastases, recent data suggest that up to one fifth are still misdiagnosed with a previously underrecognized culprit: endometrioid carcinoma. Using an index case of an ovarian endometrioid carcinoma with mucinous differentiation masquerading as a mucinous carcinoma, we sought to identify the most efficient biomarker combination that could distinguish these 2 histotypes. Eight immunohistochemical markers were assessed on tissue microarrays from 183 endometrioid carcinomas, 77 mucinous carcinomas, and 72 mucinous borderline tumors. Recursive partitioning revealed a simple 2-marker panel consisting of PR and vimentin. The combination of PR absence and vimentin absence could predict mucinous tumors with a sensitivity of 95.1%, a specificity of 96.7%, and an overall accuracy of 96.0%. Additional marker combinations did not improve accuracy. The 5-yr ovarian cancer-specific survival for mucinous carcinoma was significantly worse than endometrioid carcinoma (70% vs. 86%, respectively, P=0.02). Our proposed 2-marker algorithm allows diagnostic distinction between mucinous and endometrioid ovarian carcinomas when morphology is not straightforward. Given key differences in the underlying biology and clinical behavior of these 2 histotypes, improved diagnostic precision is essential for guiding appropriate management and treatment.

Ovarian Combined Brenner Tumor, Mucinous Cystadenoma and Struma Ovarii: First Report of a Rare Combination Brenner tumors are uncommon ovarian neoplasms which occasionally occur in combination with a mucinous tumor. Rarely, the combination of Brenner tumor and thyroid tissue (struma ovarii) has been reported. We report an ovarian neoplasm with components of Brenner tumor, mucinous cystadenoma and struma ovarii. As far as we are aware, this combination has not been previously reported. We speculate on the possible histogenesis of this combination of elements.