STK11 (LKB1) immunohistochemistry is a sensitive and specific marker for STK11 adnexal tumours.

AIMS: STK11 adnexal tumour is a rare, recently described malignant neoplasm that is associated with Peutz-Jeghers syndrome. [Correction added on 3 October 2024, after first online publication: 'ST11' in preceding sentence has been corrected to 'STK11' in this version.] It predominantly originates from the para-adnexal soft tissues and often shows secondary involvement of the fallopian tube and ovary. STK11 adnexal tumours have a broad differential diagnosis due to their variable morphology and non-specific immunoprofile, and diagnostic confirmation currently requires sequencing to identify an STK11 mutation. We investigate the diagnostic utility of STK11 (LKB1) immunohistochemistry (IHC) in a cohort of STK11 adnexal tumours and morphological mimics. METHODS AND RESULTS: IHC for STK11 was performed on 122 tumours, including 17 STK11 adnexal tumours and 105 morphological mimics (10 female adnexal tumours of Wolffian origin, 22 adult granulosa cell tumours, 10 juvenile granulosa cell tumours, four Sertoli-Leydig cell tumours, two Leydig cell tumours, one Sertoli cell tumour, one steroid cell tumour, four extra-ovarian sex cord-stromal tumours, 16 ovarian endometrioid carcinomas, eight tubo-ovarian high-grade serous carcinomas, five ovarian mesonephric-like adenocarcinomas, 14 ovarian carcinosarcomas, five peritoneal malignant mesotheliomas, two pelvic plexiform leiomyomata and one ovarian solid pseudopapillary tumour). All STK11 adnexal tumours showed complete loss of cytoplasmic staining for STK11. All other tumour types showed retained cytoplasmic staining, except for one endometrioid carcinoma with mucinous differentiation which showed complete loss of STK11 expression and a high-grade serous carcinoma with subclonal loss. CONCLUSIONS: STK11 is a highly sensitive and specific immunohistochemical marker for distinguishing STK11 adnexal tumour from its histological mimics, and can obviate the need for confirmatory molecular studies in the appropriate morphological context.

Ewing Sarcoma of the Female Genital Tract: Clinicopathologic Analysis of 21 Cases With an Emphasis on the Differential Diagnosis of Gynecologic Round Cell, Spindle, and Epithelioid Neoplasms.

Ewing sarcoma is an uncommon neoplasm considered in the differential diagnosis of tumors with "small round cell" morphology, but its occurrence in the gynecologic tract has only been sporadically documented. Herein, we describe the largest cohort of Ewing sarcoma localized to the female genital tract to date, and emphasize their clinicopathologic resemblance to more common gynecologic neoplasms. Ewing sarcoma (n=21) was retrospectively identified from 5 institutions. The average patient age was 35 (range 6-61) years. Tumor sites included uterus (n=8), cervix (n=4), vulva (n=5), vagina (n=1), broad ligament (n=1), inguinal area (n=1), and pelvis (n=1). Nine of 18 cases in which slides were available for review demonstrated only classic round cell morphology, with the remainder showing a variable combination and prominence of variant ovoid/spindle or epithelioid appearance. Tumors showed diffuse membranous reactivity for CD99 (20/20) and were positive for NKX2.2 (8/8, diffuse) and cyclin D1 (7/7, of which 3/7 were patchy/multifocal and 4/7 were diffuse). They were negative for ER (0/6) and CD10 (0/6). Three cases were initially diagnosed as endometrial stromal sarcomas. EWSR1 rearrangement was confirmed in 20/21 by fluorescence in situ hybridization (n=15) and/or sequencing (n=8). Of the eight tumors that underwent sequencing, 6 harbored FLI1 , 1 ERG, and 1 FEV as the fusion partner. Of 11 patients with available follow-up, 5 died of disease, 1 developed lung metastases and 5 are alive with no evidence of disease. Ewing sarcoma of the gynecologic tract is a rare, aggressive entity that shares some morphologic and immunohistochemical features with other more common gynecologic neoplasms. In addition to the typical round cell appearance, variant spindled/ovoid to epithelioid morphology may also be observed and should prompt consideration of this entity with appropriate immunohistochemical and/or molecular studies.

Genomic Catastrophe (Chromothripsis and Polyploidy) Correlates With Tumor Distribution in Extrauterine High-grade Serous Carcinoma.

Most extrauterine high-grade serous carcinomas (HGSCs) are thought to develop first in the distal fallopian tube. Most models of HGSC assume origin from relatively stable, noninvasive serous tubal intraepithelial carcinomas. However, widespread tumor involvement in the absence of a serous tubal intraepithelial carcinoma could occur after catastrophic genomic events (CGEs; such as chromothripsis or polyploidy). Twenty-six HGSCs assigned to fallopian tube (n = 9, group 1) and/or ovary (n = 9, group 2), and primary peritoneal (n = 8, group 3) were assessed by microarray (Oncoscan). CGEs were identified in 15/26 (57.7%); chromothripsis-like pattern in 13/26 (50.0%) and polyploidy in 6/26 (23.1%). CGE was seen in 4/9 (44.4%), 9/9 (100%), and 2/8 (25%) cases in groups 1. 2, and 3, respectively. Overall, CGEs were seen in 9/9 (100%) cases with grossly evident ovarian parenchymal involvement versus 6/17 (35.3%) without ( P = 0.0024). Ovarian size (measured on the long axis) correlated with CGE positivity ( P = 0.016). CGEs are significantly more common in HGSCs with ovarian parenchymal involvement compared with those limited to the fallopian tube and/or extraovarian tissues. These associations suggest geographically different tumor growth patterns and support the subdivision of HGSCs according to not only the stage but also tumor distribution. They have implications for clinical and pathologic presentation, trajectory of tumor evolution, and in the case of primary peritoneal HGSCs, potentially unique precursors to tumor transitions that could inform or influence cancer prevention efforts.

Prevalence and genotype distribution of human papillomavirus in cervical adenocarcinoma (usual type and variants): A systematic review and meta-analysis.

Cervical glandular neoplasms represent a heterogeneous group of tumors for which a comprehensive overview of the involvement of high-risk human papillomaviruses (HPV) in pathogenesis is still lacking. We first searched MEDLINE (PubMed), Embase, and Scopus databases (until October 2022), and systematically reviewed available literature. We then quantitatively estimated both pooled and genotype-specific prevalence of HPV DNA as well as the influence of various factors (e.g., geographical region, histological subtype, tissue/sample type) on computed effect size by means of random effects meta-analysis. In total, 379 studies comprising 17 129 cases of cervical adenocarcinoma were identified. The pooled HPV prevalence was 78.4% (95% confidence interval [95% CI]: 76.2-80.3) with a significant between-study heterogeneity (I2 = 79.4%, Q test p < 0.0001). Subgroup analyses indicated that the effect size differed substantially by geographical region (from 72.5% [95% CI: 68.7-76.1] in Asia to 86.8% [95% CI: 82.2-90.3] in Oceania) (p < 0.0001) and histological subtype of cancer (from 9.8% [95% CI: 5.5-17] in gastric-type to 85% [95% CI: 79.6-89.2] in usual-type cervical adenocarcinoma) (p < 0.0001). HPV16 and HPV18 were by far the most frequently detected viral strains with specific prevalence of 49.8% (95% CI: 46.9-52.6) and 45.3% (95% CI: 42.8-47.8), respectively. When stratified by continent or histologic variant, these genotype-specific results varied in a relatively limited manner. Altogether, these findings support that all histological subtypes of cervical adenocarcinoma are etiologically linked to high-risk HPV but to varying degrees. Therefore, a dual-criteria classification taking into account accurately both morphological and virological aspects could be an interesting evolution of the current binary World Health Organization classification, better reflecting the pathogenic diversity of the disease.

Replication stress and defective checkpoints make fallopian tube epithelial cells putative drivers of high-grade serous ovarian cancer.

Clinical and molecular evidence indicates that high-grade serous ovarian cancer (HGSOC) primarily originates from the fallopian tube, not the ovarian surface. However, the reasons for this preference remain unclear. Our study highlights significant differences between fallopian tube epithelial (FTE) and ovarian surface epithelial (OSE) cells, providing the molecular basis for FTEs as site of origin of HGSOC. FTEs, unlike OSEs, exhibit heightened replication stress (RS), impaired repair of stalled forks, ineffective G2/M checkpoint, and increased tumorigenicity. BRCA1 heterozygosity exacerbates these defects, resulting in RS suppression haploinsufficiency and an aggressive tumor phenotype. Examination of human and mouse sections reveals buildup of the RS marker 53BP1 primarily in the fallopian tubes, particularly at the fimbrial ends. Furthermore, menopausal status influences RS levels. Our study provides a mechanistic rationale for FTE as the site of origin for HGSOC, investigates the impact of BRCA1 heterozygosity, and lays the groundwork for targeting early HGSOC drivers.

Inflammatory Activity of Epithelial Stem Cell Variants from Cystic Fibrosis Lungs Is Not Resolved by CFTR Modulators.

Rationale: CFTR (cystic fibrosis transmembrane conductance regulator) modulator drugs restore function to mutant channels in patients with cystic fibrosis (CF) and lead to improvements in body mass index and lung function. Although it is anticipated that early childhood treatment with CFTR modulators will significantly delay or even prevent the onset of advanced lung disease, lung neutrophils and inflammatory cytokines remain high in patients with CF with established lung disease despite modulator therapy, underscoring the need to identify and ultimately target the sources of this inflammation in CF lungs. Objectives: To determine whether CF lungs, like chronic obstructive pulmonary disease (COPD) lungs, harbor potentially pathogenic stem cell "variants" distinct from the normal p63/Krt5 lung stem cells devoted to alveolar fates, to identify specific variants that might contribute to the inflammatory state of CF lungs, and to assess the impact of CFTR genetic complementation or CFTR modulators on the inflammatory variants identified herein. Methods: Stem cell cloning technology developed to resolve pathogenic stem cell heterogeneity in COPD and idiopathic pulmonary fibrosis lungs was applied to end-stage lungs of patients with CF (three homozygous CFTR:F508D, one CFTR F508D/L1254X; FEV1, 14-30%) undergoing therapeutic lung transplantation. Single-cell-derived clones corresponding to the six stem cell clusters resolved by single-cell RNA sequencing of these libraries were assessed by RNA sequencing and xenografting to monitor inflammation, fibrosis, and mucin secretion. The impact of CFTR activity on these variants after CFTR gene complementation or exposure to CFTR modulators was assessed by molecular and functional studies. Measurements and Main Results: End-stage CF lungs display a stem cell heterogeneity marked by five predominant variants in addition to the normal lung stem cell, of which three are proinflammatory both at the level of gene expression and their ability to drive neutrophilic inflammation in xenografts in immunodeficient mice. The proinflammatory functions of these three variants were unallayed by genetic or pharmacological restoration of CFTR activity. Conclusions: The emergence of three proinflammatory stem cell variants in CF lungs may contribute to the persistence of lung inflammation in patients with CF with advanced disease undergoing CFTR modulator therapy.

Inaugurating High-Throughput Profiling of Extracellular Vesicles for Earlier Ovarian Cancer Detection.

Detecting early cancer through liquid biopsy is challenging due to the lack of specific biomarkers for early lesions and potentially low levels of these markers. The current study systematically develops an extracellular-vesicle (EV)-based test for early detection, specifically focusing on high-grade serous ovarian carcinoma (HGSOC). The marker selection is based on emerging insights into HGSOC pathogenesis, notably that it arises from precursor lesions within the fallopian tube. This work thus establishes murine fallopian tube (mFT) cells with oncogenic mutations and performs proteomic analyses on mFT-derived EVs. The identified markers are then evaluated with an orthotopic HGSOC animal model. In serially-drawn blood of tumor-bearing mice, mFT-EV markers increase with tumor initiation, supporting their potential use in early cancer detection. A pilot clinical study (n = 51) further narrows EV markers to five candidates, EpCAM, CD24, VCAN, HE4, and TNC. The combined expression of these markers distinguishes HGSOC from non-cancer with 89% sensitivity and 93% specificity. The same markers are also effective in classifying three groups (non-cancer, early-stage HGSOC, and late-stage HGSOC). The developed approach, for the first time inaugurated in fallopian tube-derived EVs, could be a minimally invasive tool to monitor women at high risk of ovarian cancer for timely intervention.

Cloning a profibrotic stem cell variant in idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressive, irreversible, and rapidly fatal interstitial lung disease marked by the replacement of lung alveoli with dense fibrotic matrices. Although the mechanisms initiating IPF remain unclear, rare and common alleles of genes expressed in lung epithelia, combined with aging, contribute to the risk for this condition. Consistently, single-cell RNA sequencing (scRNA-seq) studies have identified lung basal cell heterogeneity in IPF that might be pathogenic. We used single-cell cloning technologies to generate "libraries" of basal stem cells from the distal lungs of 16 patients with IPF and 10 controls. We identified a major stem cell variant that was distinguished from normal stem cells by its ability to transform normal lung fibroblasts into pathogenic myofibroblasts in vitro and to activate and recruit myofibroblasts in clonal xenografts. This profibrotic stem cell variant, which was shown to preexist in low quantities in normal and even fetal lungs, expressed a broad network of genes implicated in organ fibrosis and showed overlap in gene expression with abnormal epithelial signatures identified in previously published scRNA-seq studies of IPF. Drug screens highlighted specific vulnerabilities of this profibrotic variant to inhibitors of epidermal growth factor and mammalian target of rapamycin signaling as prospective therapeutic targets. This profibrotic stem cell variant in IPF was distinct from recently identified profibrotic stem cell variants in chronic obstructive pulmonary disease and may extend the notion that inappropriate accrual of minor and preexisting stem cell variants contributes to chronic lung conditions.

Profiling extracellular vesicles in circulation enables the early detection of ovarian cancer.

Ovarian cancer is a heterogeneous group of tumors in both cell type and natural history. While outcomes are generally favorable when detected early, the most common subtype, high-grade serous carcinoma (HGSOC), typically presents at an advanced stage and portends less favorable prognoses. Its aggressive nature has thwarted early detection efforts through conventional detection methods such as serum CA125 and ultrasound screening and thus inspired the investigation of novel biomarkers. Here, we report the systematic development of an extracellular-vesicle (EV)-based test to detect early-stage HGSOC. Our study is based on emerging insights into HGSOC biology, notably that it arises from precursor lesions within the fallopian tube before traveling to ovarian and/or peritoneal surfaces. To identify HGSOC marker candidates, we established murine fallopian tube (mFT) cells with oncogenic mutations in Brca1/2, Tp53 , and Pten genes, and performed proteomic analyses on mFT EVs. The identified markers were then evaluated with an orthotopic HGSOC animal model. In serially-drawn blood samples of tumor-bearing mice, mFT-EV markers increased with tumor initiation, supporting their potential use in early cancer detection. A pilot human clinical study ( n = 51) further narrowed EV markers to five candidates, EpCAM, CD24, VCAN, HE4, and TNC. Combined expression of these markers achieved high OvCa diagnostic accuracy (cancer vs. non-cancer) with a sensitivity of 0.89 and specificity of 0.93. The same five markers were also effective in a three-group classification: non-cancer, early-stage (I & II) HGSOC, and late-stage (III & IV) HGSOC. In particular, they differentiated early-stage HGSOC from the rest with a specificity of 0.91. Minimally invasive and repeatable, this EV-based testing could be a versatile and serial tool for informing patient care and monitoring women at high risk for ovarian cancer.

Genomic catastrophe, the peritoneal cavity and ovarian cancer prevention.

The current theory of carcinogenesis for the deadliest of 'ovarian' cancers-high-grade serous carcinoma (HGSC)-holds that the malignancy develops first in the fallopian tube and spreads to the ovaries, peritoneum, and/or regional lymph nodes. This is based primarily on the observation of early forms of serous neoplasia (serous tubal intraepithelial lesions [STILs], and serous tubal intraepithelial carcinomas [STICS]) in the fimbria of women undergoing risk reduction surgery. However, these lesions are uncommon in the general population, confer a low risk (5%) of HGSC following their removal in at-risk women with germ-line BRCA1/2 mutations, and require 4 or more years to recur as intraperitoneal HGSC. These features suggest that isolated STILs and STICs behave as precursors, with uncertain cancer risk rather than carcinomas. Their evolution to HGSC within, or after, escape from the tube could proceed stepwise with multiple biologic events; however, it is unclear whether tubal or ovarian HGSCs encountered in the setting of advanced disease evolved in the same fashion. The latter scenario could also be explained by a 'catastrophic' model in which STICs suddenly develop with invasive and metastatic potential, overwhelming or obscuring the site of origin. Moreover, a similar model might explain the sudden emergence of HGSC in the peritoneal cavity following escape of precursor cells years before. Long-term follow-up data from opportunistic or prophylactic salpingectomy should shed light on where malignant transformation occurs, as well as the timeline from precursor to metastatic HGSC. © 2022 The Pathological Society of Great Britain and Ireland.

Dissection of PIK3CA Aberration for Cervical Adenocarcinoma Outcomes.

Personalized treatment of genetically stratified subgroups has the potential to improve outcomes in many malignant tumors. This study distills clinically meaningful prognostic/predictive genomic marker for cervical adenocarcinoma using signature genomic aberrations and single-point nonsynonymous mutation-specific droplet digital PCR (ddPCR). Mutations in PIK3CA E542K, E545K, or H1047R were detected in 41.7% of tumors. PIK3CA mutation detected in the patient's circulating DNA collected before treatment or during follow-up was significantly associated with decreased progression-free survival or overall survival. PIK3CA mutation in the circulating DNA during follow-up after treatment predicted recurrence with 100% sensitivity and 64.29% specificity. It is the first indication of the predictive power of PIK3CA mutations in cervical adenocarcinoma. The work contributes to the development of liquid biopsies for follow up surveillance and a possibility of tailoring management of this particular women's cancer.

Interobserver reproducibility of the diagnosis of differentiated exophytic vulvar intraepithelial lesion (DEVIL) and the distinction from its mimics.

AIMS: Most vulvar squamous cell carcinomas are human papillomavirus (HPV)-associated or TP53-mutant. A third category of HPV-independent TP53-wild-type lesions is uncommon and not fully understood. Differentiated exophytic vulvar intraepithelial lesion (DEVIL) has been characterised as a precursor of this latter category. The reproducibility of the diagnosis of DEVIL and its distinction from lesions with overlapping morphology has not been studied. Our aim was to establish the interobserver agreement in the diagnosis of DEVIL and its distinction from neoplastic and reactive conditions of the vulva on haematoxylin and eosin evaluation. METHODS AND RESULTS: A set of 35 slides was evaluated by eight reviewers (two trainees and six practising gynaecological pathologists). The set included DEVIL, condyloma, established vulvar precursors [high-grade squamous intraepithelial lesion (HSIL) and differentiated vulvar intraepithelial neoplasia (dVIN)] with superimposed acanthosis or verruciform growth, lichen simplex chronicus (LSC), and psoriasis. Kappa (κ) values were calculated. Overall, interobserver agreement was moderate (κ = 0.56), improving to substantial (κ = 0.7) when evaluation was performed by practising pathologists. Agreement was strong for the diagnosis of HSIL (κ = 0.88), and substantial for the diagnosis of DEVIL (κ = 0.61), condyloma (κ = 0.79), and LSC (κ = 0.72). Agreement was moderate for the diagnosis of dVIN (κ = 0.59) and psoriasis (κ = 0.53). Perfect agreement (6/6) among practising pathologists was observed in 43% of cases, and majority agreement (5/6 or 4/6) was observed in 48% of cases. CONCLUSIONS: Reproducibility in the diagnosis of verruciform vulvar lesions, including the novel DEVIL, is acceptable overall. Reproducibility is higher for well-known lesions such as HSIL and condyloma than for more challenging diagnoses such as DEVIL, dVIN, and psoriasis. Agreement is higher among practising gynaecological pathologists, suggesting that training and experience improve reproducibility. Our findings support the inclusion of DEVIL as a diagnostic entity in the classification of vulvar squamous lesions.

Enhanced Efficacy of Aurora Kinase Inhibitors in G2/M Checkpoint Deficient TP53 Mutant Uterine Carcinomas Is Linked to the Summation of LKB1-AKT-p53 Interactions.

Uterine carcinoma (UC) is the most common gynecologic malignancy in the United States. TP53 mutant UCs cause a disproportionate number of deaths due to limited therapies for these tumors and the lack of mechanistic understanding of their fundamental vulnerabilities. Here we sought to understand the functional and therapeutic relevance of TP53 mutations in UC. We functionally profiled targetable TP53 dependent DNA damage repair and cell cycle control pathways in a panel of TP53 mutant UC cell lines and patient-derived organoids. There were no consistent defects in DNA damage repair pathways. Rather, most models demonstrated dependence on defective G2/M cell cycle checkpoints and subsequent upregulation of Aurora kinase-LKB1-p53-AKT signaling in the setting of baseline mitotic defects. This combination makes them sensitive to Aurora kinase inhibition. Resistant lines demonstrated an intact G2/M checkpoint, and combining Aurora kinase and WEE1 inhibitors, which then push these cells through mitosis with Aurora kinase inhibitor-induced spindle defects, led to apoptosis in these cases. Overall, this work presents Aurora kinase inhibitors alone or in combination with WEE1 inhibitors as relevant mechanism driven therapies for TP53 mutant UCs. Context specific functional assessment of the G2/M checkpoint may serve as a biomarker in identifying Aurora kinase inhibitor sensitive tumors.

Mural nodules in mucinous ovarian tumors represent a morphologic spectrum of clonal neoplasms: a morphologic, immunohistochemical, and molecular analysis of 13 cases.

Mucinous ovarian tumors rarely harbor mural nodules, which have historically been classified as sarcoma-like, anaplastic carcinomatous, or sarcomatous on the basis of predominant morphologic features. The molecular relationship between mural nodules and associated mucinous ovarian tumors remains poorly characterized, as does the molecular pathogenesis of these mural nodules. Thus, we analyzed the morphological, immunohistochemical, and genetic features of 13 mucinous ovarian tumors and associated mural nodule(s). Three harbored sarcoma-like mural nodules and ten contained anaplastic carcinomatous nodules, including 1 tumor with spatially discrete anaplastic carcinomatous and sarcomatous nodules. Twelve of 13 cases showed genetic evidence of clonality between the mural nodule(s) and associated mucinous ovarian tumor, including all three tumors with sarcoma-like morphology. Mural nodules were genetically identical in the five cases in which there were multiple discrete mural nodules that were sequenced separately. MTAP and p53 immunohistochemistry confirmed the distribution of neoplastic cells in a subset of sarcoma-like and anaplastic carcinomatous nodules. No single recurrent genetic alteration was associated with mural nodule development. No recurrent genetic differences were identified between mural nodules with sarcoma-like, anaplastic carcinomatous, and sarcomatous morphology. Of 11 patients with clinical follow-up, three died of disease 3, 8, and 9 months after diagnosis, but no recurrent genetic events were associated with poor outcome. These molecular data suggest that sarcoma-like, anaplastic carcinomatous, and sarcomatous nodules represent a morphologic spectrum of clonal neoplasms arising in mucinous ovarian tumors rather than three discrete biological entities.

A Clinical and Pathologic Exploration of Suspected Peritoneal Endometriotic Lesions.

Endometriosis is generally histopathologically defined as the presence of at least 2 of the following: endometrial stroma, Müllerian epithelium, and/or hemosiderin-laden macrophages (HLM). Despite clinically evident endometriotic lesions, biopsies are frequently nondiagnostic. In this study, we conducted a large-scale review of biopsies of lesions clinically thought to represent endometriosis and correlate the histologic findings with clinical appearance to expand sensitivity of the pathologic definition of endometriosis, particularly in patients on hormonal therapy. In all, 112 biopsies from 78 patients (mean age=25, range 18-39 yr) were reviewed for histopathologic features suggestive of or diagnostic for endometriosis including the presence of endometrial stroma, Müllerian epithelium, dystrophic calcifications, HLM, chronic inflammation, adhesions, and vascular proliferation. Endometriosis was confirmed by pathologic criteria in 37 of 78 patients (47%). Biopsies from patients on hormonal therapy (n=62, 80%) were significantly less likely to meet pathologic criteria for endometriosis (P=0.01). Nondiagnostic biopsies (70/112; 63%) frequently displayed HLM (20%), chronic inflammation (29%), dystrophic calcifications (26%), vascular proliferation (20%), or adhesions (20%) and were significantly more likely to have a vascular clinical appearance (P=0.01). Diagnostic biopsies (42/112; 38%) were more likely to have a blue/black clinical appearance (P=0.03), demonstrate HLM (P=0.004), and display pseudodecidualization (P=0.05). Patients with a high clinical suspicion of endometriosis have a range of histologic findings, with less than half meeting the current histopathologic criteria for diagnosing endometriosis. Given the heterogeneous histopathologic appearance, revision of the histologic criteria may be warranted with further exploration, particularly for lesions with predominantly vascular features.

Enhanced Efficacy of Simultaneous PD-1 and PD-L1 Immune Checkpoint Blockade in High-Grade Serous Ovarian Cancer.

Immune therapies have had limited efficacy in high-grade serous ovarian cancer (HGSC), as the cellular targets and mechanism(s) of action of these agents in HGSC are unknown. Here we performed immune functional and single-cell RNA sequencing transcriptional profiling on novel HGSC organoid/immune cell co-cultures treated with a unique bispecific anti-programmed cell death protein 1 (PD-1)/programmed death-ligand 1 (PD-L1) antibody compared with monospecific anti-PD-1 or anti-PD-L1 controls. Comparing the functions of these agents across all immune cell types in real time identified key immune checkpoint blockade (ICB) targets that have eluded currently available monospecific therapies. The bispecific antibody induced superior cellular state changes in both T and natural killer (NK) cells. It uniquely induced NK cells to transition from inert to more active and cytotoxic phenotypes, implicating NK cells as a key missing component of the current ICB-induced immune response in HGSC. It also induced a subset of CD8 T cells to transition from naïve to more active and cytotoxic progenitor-exhausted phenotypes post-treatment, revealing the small, previously uncharacterized population of CD8 T cells responding to ICB in HGSC. These state changes were driven partially through bispecific antibody-induced downregulation of the bromodomain-containing protein BRD1. Small-molecule inhibition of BRD1 induced similar state changes in vitro and demonstrated efficacy in vivo, validating the co-culture results. Our results demonstrate that state changes in both NK and a subset of T cells may be critical in inducing an effective anti-tumor immune response and suggest that immune therapies able to induce such cellular state changes, such as BRD1 inhibitors, may have increased efficacy in HGSC. SIGNIFICANCE: This study indicates that increased efficacy of immune therapies in ovarian cancer is driven by state changes of NK and small subsets of CD8 T cells into active and cytotoxic states.

Synovial Sarcoma of the Female Genital Tract: A Protean Mimic of Müllerian Neoplasia.

Synovial sarcoma most commonly occurs in the extremities but has rarely been described in the female genital tract. In this series, we describe the clinical, morphologic, immunohistochemical, and molecular features of 7 cases of vulvovaginal synovial sarcoma (vulva, n=6; vagina, n=1). We emphasize their wide morphologic spectrum, which can overlap significantly with other more common tumors at these sites, as highlighted by 2 cases initially diagnosed as other entities (endometrioid carcinoma and malignant peripheral nerve sheath tumor). The average patient age was 41 (range: 23 to 62) years and tumor size ranged from 0.8 to 7 cm. Histologically, the tumors were biphasic (n=6) and monophasic (n=1). All cases were confirmed with fluorescence in situ hybridization or sequencing, and 5/5 cases were positive for the novel immunohistochemical markers SSX and SS18-SSX. In 3 cases with follow-up, 2 patients died of disease and 1 was alive with no evidence of disease. Previously described cases arising in the female genital tract are also reviewed. Vulvovaginal monophasic synovial sarcoma raises a broad differential diagnosis, including smooth muscle tumors, spindled carcinomas, and melanoma. Biphasic synovial sarcoma may mimic Müllerian carcinosarcoma, endometrioid carcinoma with spindled, corded, and hyalinized elements, and mesonephric-like adenocarcinoma. Awareness that synovial sarcoma can occur in the female genital tract with a wide variety of histologic appearances is critical for correctly diagnosing this rare entity. In particular, synovial sarcoma should be considered for any deeply situated "adenocarcinoma" in the vulva, with attention to subtle spindle cell differentiation.

Molecular characterization of diffuse malignant peritoneal mesothelioma.

Malignant peritoneal mesothelioma is a rare aggressive tumor that arises from the peritoneal lining. While recurrent BAP1 mutations have been identified in a subset of mesotheliomas, molecular characteristics of peritoneal mesotheliomas, including those lacking BAP1 alterations, remain poorly understood. Using targeted next-generation sequencing, we examined the molecular features of 26 diffuse malignant peritoneal mesotheliomas. As part of an exploratory analysis, we analyzed an additional localized peritoneal mesothelioma and one well-differentiated papillary mesothelioma with invasive foci. Genomic characterization identified categories of diffuse malignant peritoneal mesotheliomas: The first group included 18 (69%) tumors with recurrent BAP1 alterations, with eight (31%) having more than one BAP1 alterations, and concomitant alterations in PBRM1 (46%) and SETD2 (35%). All tumors with complete loss of BAP1 expression by immunohistochemistry harbored BAP1 molecular alterations. PBRM1 alterations were significantly enriched in the BAP1-altered cohort. Frequent copy number loss of BAP1, ARID1B, PRDM1, PBRM1, SETD2, NF2, and CDKN2A was noted. The second group included eight (31%) BAP1-wild-type tumors: two with TP53 mutations, one with a TRAF7 activating mutation, one with a SUZ12 inactivating mutation, and three with ALK rearrangements that we previously published. One TP53-mutant biphasic mesothelioma showed evidence of genomic near-haploidization showing loss of heterozygosity of all chromosomes except 5, 7, 16, and 20. The localized peritoneal mesothelioma harbored a nonsense CHEK2 mutation, and the well-differentiated papillary mesothelioma with invasive foci harbored no reportable variants. In conclusion, we described the genetic categories of diffuse malignant peritoneal mesotheliomas, with BAP1-mutant and BAP1-wild-type groups. Our findings implicated DNA repair, epigenetics, and cell cycle regulation in the pathogenesis of peritoneal mesotheliomas, with identification of potential therapeutic targets.

Prostatic Metaplasia of the Vagina and Uterine Cervix: An Androgen-associated Glandular Lesion of Surface Squamous Epithelium.

Prostatic-type differentiation in the lower female genital tract is encountered rarely and its causes and clinical associations are not well established. Within the vagina, reports to date have invariably described ectopic prostatic-type differentiation as restricted to the lamina propria. We recently encountered a patient receiving testosterone for gender dysphoria whose vaginectomy specimen showed a prostatic glandular proliferation within the surface epithelium. To elucidate its potential association with androgen exposure, we sought similar lesions, resected over a 26-year period, from patients with exogenous or endogenous androgen excess. Thirteen cases were identified, involving the vagina (n=12) and exocervix (n=1). The most common clinical context was gender dysphoria with long-term testosterone therapy; the lesion was present in 7 of 8 gender-dysphoric patients examined. Four other patients had congenital disorders of sexual development associated with endogenous androgen excess (congenital adrenal hyperplasia, 46,XY disorder of sexual development, and ovotesticular disorder of sexual development). Two had no known exposure to androgen excess. Immunohistochemically, glands stained for NKX3.1 (100% of cases), androgen receptor (100%), CK7 (92%), and prostate-specific antigen (69%). Follow-up (median duration, 11 mo) showed no masses or neoplasia. We propose the designation "androgen-associated prostatic metaplasia" for this form of prostate tissue with distinctive clinical, histologic and immunohistochemical features. It is novel and previously unrecognized within the vagina. It is strikingly prevalent among patients undergoing gender-affirming surgery, an increasingly common procedure. Recognition is important to distinguish it from other potentially neoplastic glandular lesions and facilitate accrual of more follow-up data to better understand its natural history.