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Fumarate hydratase tumor predisposition syndrome (FHTPS) is caused by germline fumarate hydratase (FH) pathogenic variants (PVs). Most women with FHTPS develop FH-deficient (FHD) uterine leiomyomas (ULs), which arise 10 to 15 years earlier than aggressive FHD-renal cell carcinoma. We evaluate a previously proposed FHTPS screening strategy for women with ULs. This 5-year, prospective and retrospective study performed FH and later S-(2-succino) cysteine immunohistochemistry (IHC) on all uterine smooth muscle (USM) tumors in patients 40 (later ≤30) years or younger and on all USM tumors with suggestive FHD morphology regardless of age. Patients with FHD tumors by IHC were referred to genetic counseling. Of 840 USM tumors, 112 FHD-tumors by IHC (13%) were identified, all with suggestive FHD-morphology; 44 patients (39%) underwent germline testing, and 15 harbored germline FH PVs (34.1% of germline tested, 13.4% of all FHD-tumors). While FHD tumors were seen across a wide age range (24 to 73 y), those with germline FH PVs were significantly younger (median 33 vs 44 years wild-type, P = 0.0032). Few (12.5%) patients ≥40 and no patients ≥50 had a germline FH PV, whereas a majority (60%) of patients <40 (86% of those <30) had a germline FH PV. We demonstrate that previously proposed resource-conscious screening involving morphology and IHC is effective for identifying women with FHTPS. We provide prospective data confirming patients presenting with FHD-ULs over age 50 are unlikely to harbor germline FH PVs and argue that for germline testing without consideration of other factors, a threshold of younger than 50 years may be appropriate., Competing Interests: Conflicts of Interest and Source of Funding: The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article., (Copyright © 2025 Wolters Kluwer Health, Inc. All rights reserved.)

Aims: Mesonephric-like adenocarcinoma (MLA) of the endometrium is often a diagnostic challenge, due to its morphological resemblance to other more common Müllerian neoplasms. This study aimed to retrospectively identify overlooked MLA in a large endometrial carcinoma cohort, using a combination of immunohistochemistry (IHC), morphology and KRAS sequencing., Methods and Results: IHC was conducted on 1094 endometrial carcinomas, identifying 16 potential MLA cases based on GATA3+ and/or TTF1+ and ER- staining patterns, which subsequently underwent detailed histological review, KRAS sequencing and ProMisE molecular classification. Of the IHC screen-positive cases, one was positive for both GATA3 and TTF1, nine were positive for GATA3 only and six were positive for TTF1 only. All IHC screen-positive cases were POLE wild-type. All five tumours in the NSMP category showed morphological features of MLA, while the three MMRd and eight p53abn tumours did not show MLA morphology. The five cases diagnosed as MLA on review were all originally diagnosed as low-grade endometrioid adenocarcinoma probably because of rare morphological patterns, being predominantly papillary or ductal. Four of the five cases harboured a KRAS mutation., Conclusion: This study highlights the importance of a comprehensive diagnostic approach for accurately identifying endometrial MLA and for pathologists to be aware of papillary and ductal patterns in endometrial carcinoma assessment. Further exploration into the molecular landscape of MLA is essential for refining diagnostic criteria and developing targeted therapies., (© 2024 The Author(s). Histopathology published by John Wiley & Sons Ltd.)

Aims: Decreased oestrogen receptor (ER) expression is a marker of poor prognosis in endometrial carcinomas (EC) of no specific molecular profile (NSMP), but the optimal cut-off to separate high-risk 'low ER' versus low-risk 'high ER' expression has not been defined. Here we characterised the distribution of ER staining in a cohort of ECs., Methods and Results: Biopsy specimens from 120 cases of NSMP EC were stained for ER and assigned an Allred score. In 66 additional cases ER staining of matched biopsy and hysterectomy were compared. Twelve of 120 tumours had an Allred score of 0-3, including three endometrioid carcinomas (EEA) (one G1, two G3), four clear cell carcinomas (CCC), two mesonephric-like adenocarcinoma (MLA) and one each of: gastric-type adenocarcinoma, carcinosarcoma and endometrial carcinoma NOS. Three had Allred scores of 4-5: two MLA and one high-grade carcinoma with yolk sac differentiation. Five had Allred scores of 6: four EEA (one G1, one G2, two G3) and one mixed clear cell and endometrioid carcinoma. The remaining 100 tumours with Allred scores ≥ 7 were all EEA (66 G1, 28 G2, five G3 and one grade unknown). Comparing the biopsy versus hysterectomy ER staining (n = 66), the results were within a single Allred score point, except two cases with strong diffuse expression in the biopsy (Allred 8) and moderate expression in the hysterectomy (Allred 5)., Conclusions: Most NSMP ECs (> 80%) show high ER expression (Allred score ≥ 7). All non-endometrioid carcinomas and a few endometrioid carcinomas had lower ER expression (Allred score ≤ 6) or were completely negative., (© 2024 The Author(s). Histopathology published by John Wiley & Sons Ltd.)

Summary: Our aim was to assess the molecular subtype(s) and perform a detailed morphologic review of tumors diagnosed as carcinosarcoma in a population-based cohort. Forty-one carcinosarcomas were identified from a cohort of 973 endometrial carcinomas diagnosed in 2016. We assessed immunostaining and sequencing data and undertook expert pathology reviews of these cases as well as all subsequently diagnosed (post-2016) carcinosarcomas of no specific molecular profile (NSMP) molecular subtype (n=3) from our institutions. In the 2016 cohort, 37 of the 41 carcinosarcomas (91.2%) were p53abn, 2 (4.9%) were NSMP, and 1 each (2.4%) were POLE mut and mismatch repair deficiency molecular subtypes, respectively. Of the 4 non-p53abn tumors on review, both NSMP tumors were corded and hyalinized (CHEC) pattern endometrioid carcinoma, the mismatch repair deficiency tumor was a grade 1 endometrioid carcinoma with reactive stromal proliferation, and the POLE mut tumor was grade 3 endometrioid carcinoma with spindle cell growth, that is, none were confirmed to be carcinosarcoma on review. We found 11 additional cases among the 37 p53abn tumors that were not confirmed to be carcinosarcoma on the review (3 undifferentiated or dedifferentiated carcinomas, 5 carcinomas with CHEC features, 2 carcinomas showing prominent reactive spindle cell stroma, and 1 adenosarcoma). In the review of institutional cases reported as NSMP carcinosarcoma after 2016, 3 were identified (1 adenosarcoma and 2 mesonephric-like adenocarcinoma on review). In this series, all confirmed endometrial carcinosarcomas were p53abn. The finding of any other molecular subtype in a carcinosarcoma warrants pathology review to exclude mimics., Competing Interests: The authors declare no conflict of interest., (Copyright © 2024 by the International Society of Gynecological Pathologists.)

Objectives: Ovarian carcinosarcoma (OCS) is a rare and lethal type of ovarian cancer. Despite its incredibly poor prognosis, it has received little research attention. In this study, we aim to evaluate the molecular features of OCS and elucidate their clinical significance., Study Methods: We examined 30 OCS by immunohistochemistry (IHC) and targeted panel sequencing collected from a single institution (2003-2013) as the initial molecularly characterized cohort (Cohort A). From November 2016 to April 2023, we collected an additional 67 OCS cases from three institutions across British Columbia and Alberta as the contemporary cohort (Cohort B) for clinical correlation. The Kaplan-Meier method was used to estimate overall and progression-free survival, and differences in survival rates were compared using the log-rank test. All tests were two-sided. A p -value of less than 0.05 was considered statistically significant., Results: The majority of OCS (82%) in the initial Cohort A were p53-mutated, and the carcinomatous component displayed the histological and molecular features of a high-grade tubo-ovarian serous carcinoma (HGSC-like). In a minority of OCS, the epithelial components were characteristics of endometrioid or clear cell carcinomas, and IHC staining was wild type for p53. In the contemporary Cohort B, we observed the same histological findings related to the p53 IHC staining pattern. The median overall survival of the p53-mutated HGSC-like OCS (47 patients) was significantly higher (43.5 months) compared with that of the p53 wild-type OCS (10 patients, 8.8 months; P < 0.01). Pathogenic BRCA1/2 germline/somatic mutations were observed in 7 patients (17.5%) of HGSC-like OCS, and all these patients were alive at 3 years from diagnosis compared to a 51% 3-year survival among the patients with BRCA1/2 wild-type HGSC-like OCS (33 patients) ( p = 0.022). Majority of patients (6/7) with BRCA1/2 -mutated OCS received poly (ADP-ribose) polymerase inhibitor as maintenance therapy in this cohort., Conclusions: Most OCSs have a morphologic and molecular profile resembling HGSC; however, some OCSs display a molecular profile that suggests origin through non-serous oncogenic pathways. This molecular distinction has both prognostic and treatment (predictive) implications. These findings underscore the importance of routine p53 IHC testing on all OCS and BRCA1/2 testing on p53-mutated OCS., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Dhillon, Llaurado-Fernandez, Tessier-Cloutier, Sy, Bassiouny, Han, Wong, McRae, Kinloch, Pors, Hopkins, Covens, Köbel, Lee and Carey.)

Dedifferentiated and undifferentiated ovarian carcinomas (DDOC/UDOC) are rare neoplasms defined by the presence of an undifferentiated carcinoma. In this study, we detailed the clinical, pathological, immunohistochemical, and molecular features of a series of DDOC/UDOC. We collected a multi-institutional cohort of 23 DDOC/UDOC and performed immunohistochemistry for core switch/sucrose nonfermentable (SWI/SNF) complex proteins (ARID1A, ARID1B, SMARCA4, and SMARCB1), mismatch repair (MMR) proteins, and p53. Array-based genome-wide DNA methylation and copy number variation analyses were performed on a subset of cases with comparison made to a previously reported cohort of undifferentiated endometrial carcinoma (UDEC), small cell carcinoma of the ovary, hypercalcemic type (SCCOHT), and tubo-ovarian high-grade serous carcinoma (HGSC). The age of all 23 patients with DDOC/UDOC ranged between 22 and 71 years (with an average age of 50 years), and a majority of them presented with extraovarian disease (16/23). Clinical follow-up was available for 19 patients. Except for 2 patients, the remaining 17 patients died from disease, with rapid disease progression resulting in mortality within a year in stage II-IV settings (median disease-specific survival of 3 months). Eighteen of 22 cases with interpretable immunohistochemistry results showed loss of expression of core SWI/SNF protein(s) that are expected to result in SWI/SNF complex inactivation as 10 exhibited coloss of ARID1A and ARID1B, 7 loss of SMARCA4, and 1 loss of SMARCB1. Six of 23 cases were MMR-deficient. Two of 20 cases exhibited mutation-type p53 immunoreactivity. Methylation profiles showed coclustering of DDOC/UDOC with UDEC, which collectively were distinct from SCCOHT and HGSC. However, DDOC/UDOC showed an intermediate degree of copy number variation, which was slightly greater, compared with SCCOHT but much less compared with HGSC. Overall, DDOC/UDOC, like its endometrial counterpart, is highly aggressive and is characterized by frequent inactivation of core SWI/SNF complex proteins and MMR deficiency. Its molecular profile overlaps with UDEC while being distinct from SCCOHT and HGSC., (Copyright © 2023 United States & Canadian Academy of Pathology. Published by Elsevier Inc. All rights reserved.)

We present the clinicopathologic and immunohistochemical features of 14 endometrial glandular proliferations with conspicuous corded and hyalinized (CH) features comprised entirely or predominantly of endometrial hyperplasia. Endometrial glandular lesions ranged in severity from endometrial hyperplasia with and without cytologic atypia (5/14 [36%]) to hyperplasia with architectural complexity bordering on well-differentiated endometrioid adenocarcinoma (3/14 [21%]) to frank corded and hyalinized endometrial carcinoma ("CHEC") (6/14 [43%]). In addition to sex cord-like growth and hyalinized stroma, other common histologic features included prominent spindle cells (11/14 [79%]), keratinizing and/or morular squamous differentiation (10/14 [71%]), and osseous metaplasia (6/14 [43%]). Immunohistochemical characterization revealed aberrant nuclear beta-catenin in all cases (14/14 [100%]); additionally, all cases demonstrated positive estrogen receptors, intact PTEN, PMS2 and MSH6, and wild-type p53 expression. Patients ranged in age from 24 to 58 (mean 38) years. Of 5 patients with hyperplasia with CH features, 2 experienced complete resolution after progestin therapy and none progressed to adenocarcinoma (mean follow-up 15.6 mo, range 2 to 64). By contrast, of 2 patients with hyperplasia bordering on CHEC and with available follow-up, both subsequently developed adenocarcinoma, suggesting that even focal increased architectural complexity may predict an elevated risk of malignancy. We conclude that CH morphology is not limited to endometrioid carcinoma and may occur across a spectrum of neoplastic proliferations, including those without sufficient architectural complexity or cytologic atypia to warrant classification as adenocarcinoma. We propose the term "corded and hyalinized endometrial hyperplasia" to describe this precursor lesion and report favorable outcomes with conservative treatment., Competing Interests: Conflicts of Interest and Source of Funding: The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Uroplakin II (UPII) has been shown as a highly specific marker of urothelial carcinoma; however, it can also stain subtypes of apocrine-differentiated breast carcinoma. Given that urothelium and breast epithelium share other common immunohistochemical markers, such as CK7 and GATA3, this can lead to a potential diagnostic pitfall. We stained a cohort of triple-negative breast cancer with UPII. Compared with the diffuse, cytoplasmic staining in urothelial carcinoma, UPII was positive in 38.9% of apocrine carcinoma (7/18) with a course, granular cytoplasmic staining pattern and negative in all nonapocrine triple-negative breast cancer cases. Furthermore, the same staining pattern was present in all apocrine metaplasia of the breast (4/4) and apocrine sweat glands in normal skin (6/6). This distinct subcellular localization of UPII staining in breast carcinoma can offer a potential solution to the above diagnostic pitfall., Competing Interests: The authors declare no conflicts of interests., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

The comprehensive genomic analysis of endometrial carcinoma (EC) by The Cancer Genome Atlas (TCGA) led to the discovery of four distinct and prognostically significant molecular subgroups. Molecular classification has the potential to improve risk-stratification when integrated with clinicopathologic features and has recently been included in national and international patient management EC guidelines. Thus, the adoption of molecular classification into routine pathologic and clinical practice is likely to grow significantly in the upcoming years. Establishing an efficient and standardized workflow for performing molecular classification on ECs, and reporting both the molecular and histologic findings in an integrative manner, is imperative. Here we describe our effort to implement rapid and routine molecular classification on all ECs diagnosed at our institution. To this effect, we performed immunohistochemistry as a surrogate marker for identifying genetic and/or epigenetic alterations in DNA mismatch repair (e.g., MLH1, PMS2, MSH6, MSH2), and TP53 genes. In addition, we have developed and employed a single-gene POLE SNaPshot assay, which is a rapid and analytically sensitive method for detecting select POLE exonuclease domain mutations (EDMs). We report our molecular testing workflow and integrative reporting system as well as the clinicopathologic and molecular features of 310 ECs that underwent routine molecular classification at our institution. The 310 ECs were molecularly classified as follows: 15 (5%) POLE mutant (POLEmut), 79 (25%) mismatch repair-deficient (MMRd), 135 (44%) no specific molecular profile (NSMP), and 81 (26%) p53 abnormal (p53abnl). This work provides an initial framework for implementing routine molecular classification of ECs., (© 2021. The Author(s), under exclusive licence to United States & Canadian Academy of Pathology.)

Fumarate hydratase-deficient leiomyomas (dFH leiomyomas) often display atypical pathologic features yet exhibit a benign clinical course. Recent data suggest that dFH leiomyomas may be misclassified as smooth muscle tumors of uncertain malignant potential, a category that encompasses a heterogenous subgroup of uterine neoplasms with smooth muscle differentiation and atypical features that impart ambiguity regarding their expected clinical behavior. dFH leiomyomas can be seen in the context of hereditary leiomyomatosis and renal cell carcinoma syndrome or in the sporadic setting. In this retrospective study, we sought to examine the prevalence and clinicopathologic characteristics of dFH leiomyomas in 48 tumors previously diagnosed as smooth muscle tumors of uncertain malignant potential from 38 patients. Of these 48 tumors, 3 (6.3%) occurring in 2 patients were found to be deficient for FH by immunohistochemistry, including 1 uterine and 2 extrauterine (abdominopelvic) tumors. The 3 tumors showed histologic features typical of dFH leiomyomas, including hemangiopericytoma-like vessels, edema, macronucleoli, and atypia. Neither patient developed recurrent leiomyomas or renal cell carcinoma, and both were alive without disease at last follow-up. Our data suggest that dFH leiomyomas should be considered in the differential diagnosis of smooth muscle tumors of uncertain malignant potential, even in the context of extrauterine disease. Identification of FH deficiency in these tumors supports their classification as dFH leiomyomas despite their atypical morphologic features and/or clinical presentation. Importantly, detection of dFH in these cases may identify women at increased risk for hereditary leiomyomatosis and renal cell carcinoma who would benefit from genetic counseling and consideration for FH germline testing., Competing Interests: The authors declare no conflict of interest., (Copyright © 2021 by the International Society of Gynecological Pathologists.)

Background Observations of first trimester human trisomy 15 (T15) embryos have identified meromelic changes in the upper limbs. These changes are similar to those observed in animal studies investigating the effects of overexpression of Meis2 , a signaling transcription factor expressed during forelimb development. Although it would be exceedingly difficult to assess MEIS2 expression in the human embryonic arm, MEIS2 has been reported as consistently expressed in first trimester placental villus stroma. Methods: This study addresses whether gene dosage effect might underlie meromelia in T15 by comparing MEIS2 expression in placentas from T15 and euploid spontaneous abortions employing manual and automated assessment of MEIS2 immunohistochemical scoring. Results: Average MEIS2 expression is increased in T15 first trimester placental tissue compared to euploid controls but that the difference is marginal. Manual and automated scoring showed moderately strong correlation. Conclusion: Extrapolation of these results suggests MEIS2 overexpression may not be required for meromelia in T15.

Recently, the International Endocervical Adenocarcinoma Criteria and Classification (IECC) has reorganized the classification of endocervical adenocarcinomas (ECAs), separating them into human papilloma virus (HPV)-associated (HPVA) and HPVA independent (HPVI) categories. In this study, we sought to revalidate the IECC clinical findings in an independent cohort and assess the mutational differences between HPVA and HPVI ECAs using next generation sequencing. Consecutive cases of ECAs were reclassified under the IECC. Clinicopathologic information was collected and tissue was sent for targeted next-generation sequencing in 33 genes. Associations between HPV status, clinicopathologic parameters and mutation status, with survival were evaluated. The series comprised of 85/100 HPVA (63 HPVA-usual type, 4 villoglandular, 3 mucinous intestinal, 15 mucinous not otherwise specified) and 15/100 HPVI (9 gastric, 4 mesonephric, 1 clear cell, 1 not otherwise specified). HPVA ECAs presented at a lower age (P=0.001), smaller tumor sizes (P=0.011), less margin positivity (P=0.027), less Silva pattern C (P=0.002), and lower FIGO stages (P=0.020). HPVA had superior survival compared with HPVI ECA [overall survival (P=0.0026), disease-specific survival (P=0.0092), and progression-free survival (P=0.0041)]. Factors that correlated with worse prognosis irrespective of HPV status were FIGO stage, positive margins and lymphovascular invasion (P<0.05). TP53 mutations were detected in a significantly higher proportion of HPVIs than HPVAs (P<<0.001). The study revalidates the IECC system by reaffirming the clinical and prognostic differences between HPVA and HPVI ECAs in an independent dataset., Competing Interests: The authors declare no conflict of interest., (Copyright © 2021 by the International Society of Gynecological Pathologists.)

Recurrent vulvar squamous cell carcinomas (SCCs) are a poorly understood and aggressive group of treatment-resistant neoplasms. Currently, it remains unclear whether these are in fact recurrences of the same primary tumor, or the development of entirely new tumors. Here, to address this question, we examined the mutational profile of a series of patients with recurrent or multifocal non-human papilloma virus (HPV)-associated vulvar SCC. We performed a targeted 33-gene next-generation sequencing panel on a series of 14 patients with recurrent or multifocal non-HPV-associated vulvar SCC and precursor neoplasms. This amounted to 54 cases (33 SCC, 1 verrucous carcinoma, 13 differentiated vulvar intraepithelial neoplasia, and 7 differentiated exophytic vulvar intraepithelial lesion), with 79 mutations detected altogether. TP53 [51/79 (65%)] was the most frequently mutated gene. Mutations in PIK3CA [16/79 (20%)), HRAS [6/79 (8%)], PTEN [4/79 (5%)], EGFR [1/79 (1%)], and GNAS [1/79 (1%)] were occasionally seen. Most patients with SCC [5/9 (56%)] recurrent, 4/5 (80%) multifocal] demonstrated a clonal relationship, and harbored the same mutations in the same genes in metachronous or synchronous tumors. A subset of the recurrent tumors [2/5 (40%)] recurred with additional mutations. These clonal relationships were shared between SCC and differentiated vulvar intraepithelial neoplasia in each case. By contrast, a small number of recurrent tumors [3/9 (33%)] demonstrated novel mutations, entirely different from the primary tumor. Thus, our findings suggest that recurrent non-HPV-associated vulvar SCC can arise from 2 mechanisms., Competing Interests: The authors declare no conflict of interest., (Copyright © 2020 by the International Society of Gynecological Pathologists.)

Endometrial carcinoma (EC) is classified into a wide range of morphological variants; this list has expanded over the past decade with the inclusion of mesonephric-like and dedifferentiated carcinoma as EC variants in the fifth edition of the WHO Classification of Female Genital Tumours, and recognition that carcinosarcoma is a biphasic carcinoma rather than a sarcoma. Each EC variant has distinct molecular abnormalities, including TCGA-based molecular subtypes, allowing further subclassification and adding complexity. In contrast to this rapid progress in understanding EC, there are only two recognized EC precursor lesions: endometrial atypical hyperplasia/endometrioid intraepithelial neoplasia (EAH/EIN) and serous intraepithelial carcinoma, a situation that has not changed for many years. Diagnosis of EC precursors is a cornerstone of surgical pathology practice, with early diagnosis contributing to the relatively favorable prognosis of EC. In this review we relate the precursor lesions to each of the EC morphological variants and molecular subtypes, discuss how successful early diagnosis is for each variant/molecular subtype and how it might be improved, and identify knowledge gaps where there is insufficient understanding of EC histogenesis. © 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd., (© 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.)

Mesonephric adenocarcinoma (MA) and mesonephric-like adenocarcinoma (MLA) are uncommon neoplasms of the gynecologic tract that have until recently been poorly understood. Although their morphologic, immunohistochemical, and molecular profiles have been recently defined, little is known about their clinical behavior. Small studies have demonstrated inconsistent findings and no large studies have examined the clinical behavior of these adenocarcinomas. In this multi-institutional study, representing the largest and most stringently defined cohort of cases to date, we examined the clinicopathologic features of 99 MAs and MLAs (30 MAs of the uterine cervix, 44 MLAs of the endometrium, and 25 MLAs of the ovary). Only tumors with characteristic mesonephric morphology and either immunohistochemical or molecular support were included. Our results demonstrate that the majority of mesonephric neoplasms presented at an advanced stage (II to IV) (15/25 [60%] MA of the cervix, 25/43 [58%] MLA of the endometrium, and 7/18 [39%] MLA of the ovary). The majority (46/89 [52%] overall, 12/24 [50%] MA of the cervix, 24/41 [59%] MLA of the endometrium, and 10/24 [42%] MLA of the ovary) developed recurrences, most commonly distant (9/12 [75%] MA of the cervix, 22/24 [92%] MLA of the endometrium, and 5/9 [56%] MLA of the ovary). The 5-year disease-specific survival was 74% (n=26) for MA of cervix, 72% (n=43) for MLA of endometrium, and 71% (n=23) for MLA of ovary. Our results confirm that mesonephric neoplasms are a clinically aggressive group of gynecologic carcinomas that typically present at an advanced stage, with a predilection for pulmonary recurrence., Competing Interests: Conflicts of Interest and Source of Funding: The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

Mesonephric carcinomas (MEs) and female adnexal tumors of probable Wolffian origin (FATWO) are derived from embryologic remnants of Wolffian/mesonephric ducts. Mesonephric-like carcinomas (MLCs) show identical morphology to ME of the cervix but occur in the uterus and ovary without convincing mesonephric remnants. ME, MLC, and FATWO are challenging to diagnose due to their morphologic similarities to Müllerian/paramesonephric tumors, contributing to a lack of evidence-based and tumor-specific treatments. We performed whole-proteomic analysis on 9 ME/MLC and 56 endometrial carcinomas (ECs) to identify potential diagnostic biomarkers. Although there were no convincing differences between ME and MLC, 543 proteins showed increased expression in ME/MLC relative to EC. From these proteins, euchromatic histone lysine methyltransferase 2 (EHMT2), glutathione S-transferase Mu 3 (GSTM3), eukaryotic translation elongation factor 1 alpha 2 (EEF1A2), and glycogen synthase kinase 3 beta were identified as putative biomarkers. Immunohistochemistry was performed on these candidates and GATA3 in 14 ME/MLC, 8 FATWO, 155 EC, and normal tissues. Of the candidates, only GATA3 and EHMT2 were highly expressed in mesonephric remnants and mesonephric-derived male tissues. GATA3 had the highest sensitivity and specificity for ME/MLC versus EC (93% and 99%) but was absent in FATWO. EHMT2 was 100% sensitive for ME/MLC & FATWO but was not specific (65%). Similarly, EEF1A2 was reasonably sensitive to ME/MLC (92%) and FATWO (88%) but was the least specific (38%). GSTM3 performed intermediately (sensitivity for ME/MLC and FATWO: 83% and 38%, respectively; specificity 67%). Although GATA3 remained the best diagnostic biomarker for ME/MLC, we have identified EHMT2, EEF1A2, and GSTM3 as proteins of interest in these cancers. FATWO's cell of origin is uncertain and remains an area for future research., (Crown Copyright © 2020. Published by Elsevier Inc. All rights reserved.)

Human papillomavirus (HPV)-independent vulvar squamous cell carcinoma (VSCC) is an aggressive clinical entity. Current diagnostic guidelines for premalignant lesions are ambiguous, and their molecular profile and progression events are still unclear. We selected 75 samples, from 40 patients, including 33 VSCC, 8 verrucous carcinomas (VC), 13 differentiated-type vulvar intraepithelial neoplasia (dVIN), 11 suspicious for dVIN (?dVIN), 6 differentiated exophytic vulvar intraepithelial lesions (DE-VIL), 2 vulvar acanthosis with altered differentiation (VAAD), and 2 usual-type vulvar intraepithelial neoplasia (uVIN/HSIL). Invasive and precursor lesions were matched in 29 cases. Clinical information, p16 immunohistochemistry, and mutation analysis were performed on all lesions. All dVIN, ?dVIN, DE-VIL, and VAAD were p16 negative, all uVIN/HSIL were p16 positive. In the HPV-independent group, mutations were identified in 6 genes: TP53 (n = 40), PIK3CA (n = 20), HRAS (n = 12), MET (n = 5), PTEN (n = 4), and BRAF (n = 1). TP53 mutations occurred in 73% (22/30) VSCC, 85% (11/13) dVIN, 70% (7/10) ?dVIN and no VC (0/8), DE-VIL (0/6) nor VAAD (0/2). Basal atypia was the only reliable feature of TP53 mutations. ?dVIN lesions that were non-acanthotic and atypical but obscured by inflammation, all harbored TP53 mutations. In lesions without TP53 mutations, PIK3CA (50% VC, 33% DE-VIL, 100% VAAD, 40% VSCC) and HRAS (63% VC, 33% DE-VIL, 0% VAAD, 20% VSCC) mutations were found. Mutational progression from in situ to invasive was seen (7/26, 27%) and usually involved TP53 (4/26, 15%). Cases with TP53 and PIK3CA co-mutations had the worse clinical outcomes (p < 0.001). We recommend testing for p53 in all HPV-independent lesions suspicious for dVIN, even in the presence of marked inflammation or non-acanthotic skin, particularly when close to a margin. VC, VAAD, and DE-VIL, were almost never mutated for TP53, but instead often harbored PIK3CA and HRAS mutations. In VSCC, combined TP53 and PIK3CA mutations may inform prognosis.

Background: The International Endocervical Adenocarcinoma Criteria and Classification (IECC) separated endocervical adenocarcinomas into human papillomavirus (HPV) associated (HPVA) and non-HPV-associated (NHPVA) categories by morphology alone. Our primary objective was to assess the accuracy of HPV prediction by the IECC system compared to p16 immunohistochemistry and HPV RNA in-situ hybridization (RISH). Our secondary goal was to directly compare p16 and HPV RISH concordance., Methods: Cases were classified by IECC and stained for p16 and HPV RISH on tissue microarray, with discordant p16/HPV RISH cases re-stained on whole tissue sections. Remaining discordant cases (p16/HPV, IECC/p16, IECC/HPV discordances) were re-reviewed by the original pathologists (n = 3) and external expert pathologists (n = 2) blinded to the p16 and HPV RISH results. Final IECC diagnosis was assigned upon independent agreement between all reviewers., Results: One hundred and eleven endocervical adenocarcinomas were classified originally into 94 HPVA and 17 NHPVA cases. p16 and HPV RISH was concordant in 108/111 cases (97%) independent of the IECC. HPV RISH and p16 was concordant with IECC in 103/111 (93%) and 106/111 (95%), respectively. After expert review, concordance improved to 107/111 (96%) for HPV RISH. After review of the eight discordant cases, one remained as HPVA, four were reclassified to NHPVA from HPVA, two were unclassifiable, and one possibly represented a mixed usual and gastric-type adenocarcinoma., Conclusions: p16 and HPV RISH have excellent concordance in endocervical adenocarcinomas, and IECC can predict HPV status in most cases. Focal apical mitoses and apoptotic debris on original review led to the misclassification of several NHPVA as HPVA.

We have recently encountered p53 immunohistochemical (IHC) patterns in human papillomavirus (HPV)-associated carcinomas of the gynecologic tract, which were confused with absent (null) or overexpression TP53 mutational staining. We therefore evaluated p53 and p16 IHC in 25 squamous cell carcinomas (SCC) (16 vulva, 4 Bartholin's gland, and 5 cervix), 20 endocervical adenocarcinomas (EDAC), 14 high-grade squamous intraepithelial lesions (HSIL), 2 adenocarcinoma in situ (AIS), all of which exhibited morphologic features of HPV. Only cases showing diffuse/strong block-like p16 staining were included for further study. All EDACs underwent TP53 sequencing and HPV in situ hybridization (ISH) was performed in selected cases. p53 IHC staining fell into two main patterns. The most common was designated as "markedly reduced (null-like)" (absence or significantly attenuated staining in >70% of cells), which could be confused with true null mutational pattern. This was present in 14/25 (56%) SCCs, 7/14 (50%) HSILs, and 18/20 (90%) EDACs. The second notable pattern was "mid-epithelial (basal sparing)" (distinct absence of staining in basal cells juxtaposed with strong staining in parabasal cells), seen in 10/25 (40%) SCC, 7/14 (50%) HSIL, and none of the EDACs. There was scattered weak to moderate p53 staining (conventional wild type) in 1/25 (4%) SCC and 2/20 (10%) EDAC. No cases showed strong/diffuse overexpression. One EDAC had a TP53 missense mutation and exhibited "markedly reduced (null-like)" staining. HPV ISH revealed an inverse relationship with p53, cells positive for HPV mRNA were negative for p53. Knowledge of these patterns can help pathologists avoid misinterpreting p53 status in the setting of HPVA cancers.

Mesonephric carcinoma is a rare gynecologic neoplasm commonly mistaken for clear cell carcinoma, because of their overlapping morphologic features. Both tumors are negative for estrogen receptor and p16, magnifying this diagnostic dilemma. Recently, hepatocyte nuclear factor-1 beta (HNF-1β), a marker for clear cell carcinoma, has also been shown to be positive in mesonephric carcinomas. Other more recent markers for clear cell carcinoma, however, such as Napsin-A and alpha-methylacyl-CoA racemase (AMACR), have not yet been studied in mesonephric carcinomas. Here we examine HNF-1β, AMACR, and Napsin-A immunohistochemistry in 18 mesonephric and 55 endometrial/cervical clear cell carcinomas. HNF-1β was considered positive if nuclear staining was present in ≥70% of cells and at least moderate intensity; for Napsin-A and AMACR, any cytoplasmic staining was considered positive (≥1%). H-scores were determined by multiplying the intensity score by proportion score. HNF-1β was positive in a substantial portion of mesonephric carcinomas (9/18, 50%; H-score 98) and clear cell carcinomas (34/55, 62%; H-score 163) and did not distinguish between the 2 entities (specificity, 50%; P-value of H-score=0.08). Napsin-A and AMACR expression was significantly higher in clear cell [43/55 (78%) and 41/55 (75%), respectively] than mesonephric carcinomas [4/18 (22%) and 4/18 (22%) respectively], and helpful in this differential (specificity: 78% and 78%; P<0.05 for both). When Napsin-A and AMACR staining were seen in mesonephric carcinomas, staining was focal (≤5%), whereas staining in clear cell carcinomas was patchy/diffuse. In summary, Napsin-A and AMACR are helpful in distinguishing mesonephric carcinomas from clear cell carcinomas of the female genital tract, but HNF-1β is not.

The recent literature has shown that vulvar squamous cell carcinoma (VSCC) can be stratified into two prognostically relevant groups based on human papillomavirus (HPV) status. The prognostic value of p53 for further sub-stratification, particularly in the HPV-independent group, has not been agreed upon. This disagreement is likely due to tremendous variations in p53 immunohistochemical (IHC) interpretation. To address this problem, we sought to compare p53 IHC patterns with TP53 mutation status. We studied 61 VSCC (48 conventional VSCC, 2 VSCC with sarcomatoid features, and 11 verrucous carcinomas) and 42 in situ lesions (30 differentiated vulvar intraepithelial neoplasia [dVIN], 9 differentiated exophytic vulvar intraepithelial lesions [deVIL], and 3 high-grade squamous intraepithelial lesions or usual vulvar intraepithelial neoplasia [HSIL/uVIN]). IHC for p16 and p53, and sequencing of TP53 exons 4-9 were performed. HPV in situ hybridization (ISH) was performed in selected cases. We identified six major p53 IHC patterns, two wild-type patterns: (1) scattered, (2) mid-epithelial expression (with basal sparing), and four mutant patterns: (3) basal overexpression, (4) parabasal/diffuse overexpression, (5) absent, and (6) cytoplasmic expression. These IHC patterns were consistent with TP53 mutation status in 58/61 (95%) VSCC and 39/42 (93%) in situ lesions. Cases that exhibited strong scattered staining and those with a weak basal overexpression pattern could be easily confused. The mid-epithelial pattern was exclusively observed in p16-positive lesions; the basal and parabasal layers that had absent p53 staining, appeared to correlate with the cells that were positive for HPV-ISH. This study describes a pattern-based p53 IHC interpretation framework, which can be utilized as a surrogate marker for TP53 mutational status in both VSCC and vulvar in situ lesions.

Mesonephric carcinoma is a rare malignancy, thought to derive from Wolffian remnants. To date, no targeted molecular therapeutic options have been identified. On the basis of limited case reports, c-KIT immunohistochemical expression has been reported in female adnexal tumors of Wolffian origin, and targeted therapy with Imatinib has been attempted with mixed success. Currently, it is unclear whether c-KIT immunohistochemical expression is seen in mesonephric carcinoma, a tumor that is thought to be related to female adnexal tumors of Wolffian origin, and how this correlates with KIT mutational status. In this study, we assessed the immunohistochemical expression of c-KIT and KIT mutational status, in a series of 13 mesonephric neoplasms (5 cervical [including 2 cervical carcinosarcomas], 3 uterine corpora, 4 ovarian, and 1 vaginal/pelvic). The intensity of staining and proportion of cells showing cytoplasmic/membranous staining for c-KIT were recorded. KIT was sequenced using a next-generation sequencing panel that targeted 120 hotspots and 17 exons in 33 known actionable cancer genes. This panel included KIT exons 9, 11, and 13, and 6 hotspots (T670, D816, D820, N822, Y823, A829). Although c-KIT immunohistochemical expression was observed in the majority of mesonephric carcinomas (10/12 cases; 83%), no KIT mutations were detected. This cautions pathologists against the use of c-KIT immunohistochemistry as a surrogate marker for KIT-activating mutations in this setting. Consistent with previous studies, the majority of mesonephric neoplasms (10/13; 77%) harbored KRAS mutations. Additional mutations were found in CTNNB1 (2/13, 15%), TP53 (2/13, 15%), and PIK3CA (1/13, 8%).

The separation of benign from malignant mesothelial proliferations is a morphologically difficult problem. Mutations/deletions of components of the Hippo pathway are frequent in malignant mesotheliomas, and one downstream effect of aberrant Hippo signaling is increased production of cyclin D1. We examined expression of cyclin D1 nuclear staining in two tissue microarrays containing 52 reactive epithelial mesothelial proliferations, 51 reactive spindle cell mesothelial proliferations, 54 epithelial mesotheliomas, and 22 sarcomatous/desmoplastic mesotheliomas. When present, cyclin D1 staining was always strong, hence the arrays were scored as 0, 1-25%, 26-50%, 51-75%, and 76-100% staining. Both arrays showed a similar pattern. Reactive epithelial proliferations generally showed no staining (42/52 cases) or 1-25% staining (10/52 cases) with no cases showing >25% staining. Overall for reactive epithelial proliferations the maximum staining was 14.8% and mean 1.1 ± 2.9%. For epithelial mesotheliomas 39/54 (72%) cases demonstrated >25% staining, with 8/54 in the 26-50% staining range, 9/54 in the 51-75% range, and 22/54 in the >75% range. Combinations of staining using cyclin D1 >50% plus BAP1 or MTAP loss in epithelial mesotheliomas produced about a 10% increase in sensitivity. Reactive spindle cell proliferations showed a broader range of staining with 27/51 in the 1-25% range, 5/51 in the 26-50% range, and 1/51 >50%. Eleven of 22 sarcomatous/desmoplastic mesotheliomas scored 50% or greater. We conclude that for epithelial mesothelial proliferations, the finding of >50% of tumor cells staining supports a diagnosis of epithelial mesothelioma with 100% specificity but only modest (57%) sensitivity.

Mesonephric carcinomas of the gynecologic tract are neoplasms that are often under-recognized due to their varied morphologic appearances. Recently, GATA3 and TTF1 have been reported to be useful immunohistochemical markers for distinguishing mesonephric carcinomas from its morphologic mimics. Herein, we compared the performance of GATA3 and TTF1 to the traditional markers used for mesonephric carcinomas, CD10 and calretinin. We studied 694 cases: 8 mesonephric carcinomas (7 cervical [includes 3 mesonephric carcinosarcomas], 1 vaginal), 5 mesonephric-like carcinomas (4 uterine corpus, 1 ovarian), 585 endometrial adenocarcinomas, and 96 cervical adenocarcinomas. Mesonephric-like carcinomas were defined as tumors exhibiting the classic morphologic features of mesonephric carcinoma, but occurring outside of the cervix and without convincing mesonephric remnants. GATA3 had the highest sensitivity and specificity (91% and 94%) compared with TTF1 (45% and 99%), CD10 (73% and 83%), and calretinin (36% and 89%). GATA3, however, also stained a substantial number of uterine carcinosarcomas (23/113, 20%). TTF1 was positive in 5/5 (100%) mesonephric-like carcinomas and only 1/8 (13%) mesonephric carcinomas. In 4/6 (67%) TTF1 positive cases, GATA3 exhibited an inverse staining pattern with TTF1. In summary, GATA3 was the best overall marker for mesonephric and mesonephric-like carcinomas, but cannot be used to distinguish mesonephric carcinosarcomas from Müllerian carcinosarcomas. The inverse staining pattern between GATA3 and TTF1, suggests that TTF1 may be useful when GATA3 is negative in small biopsies where mesonephric or mesonephric-like carcinoma is suspected. The greater TTF1 positivity in mesonephric-like carcinomas suggests they may be biologically different from prototypical mesonephric carcinomas.

After acquisition, hippocampus-dependent memories undergo a systems consolidation process, during which they become independent of the hippocampus and dependent on the anterior cingulate cortex (ACC) for memory expression. However, consolidated remote memories can become transiently hippocampus-dependent again following memory reactivation. How this systems reconsolidation affects the role of the ACC in remote memory expression is not known. Using contextual fear conditioning, we show that the expression of 30-day-old remote memory can transiently be supported by either the ACC or the dorsal hippocampus following memory reactivation, and that the ACC specifically mediates expression of remote generalized contextual fear memory. We found that suppression of neural activity in the ACC with the AMPA/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) impaired the expression of remote, but not recent, contextual fear memory. Fear expression was not affected by this treatment if preceded by memory reactivation 6 h earlier, nor was it affected by suppression of neural activity in the dorsal hippocampus with the GABA-receptor agonist muscimol. However, simultaneous targeting of both the ACC and the dorsal hippocampus 6 h after memory reactivation disrupted contextual fear memory expression. Second, we observed that expression of a 30-day-old generalized contextual fear memory in a novel context was not affected by memory reactivation 6 h earlier. However, intra-ACC CNQX infusion before testing impaired contextual fear expression in the novel context, but not the original training context. Together, these data suggest that although the dorsal hippocampus may be recruited during systems reconsolidation, the ACC remains necessary for the expression of generalized contextual fear memory.

We introduce the concept of Shannon dimensionality D as a new way to quantify bipartite entanglement as measured in an experiment. This is applied to orbital-angular-momentum entanglement of two photons, using two state analyzers composed of a rotatable angular-sector phase plate that is lens coupled to a single-mode fiber. We can deduce the value of D directly from the observed two-photon coincidence fringe. In our experiment, D varies between 2 and 6, depending on the experimental conditions. We predict how the Shannon dimensionality evolves when the number of angular sectors imprinted in the phase plate is increased and anticipate that D approximately 50 is experimentally within reach.

We present a simple method to determine the cutting angle and thickness of birefringent crystals. Our method is based on chromatic polarization interferometry and allows for accuracies of typically 0.1 degrees in the cutting angle and 0.5% in the thickness.