Molecular classification of endometrial tumors is now incorporated into the 2023 FIGO staging criteria. The POLE-mutated (POLEmut) subtype is reported to have a favorable prognosis despite aggressive morphologic features. Our institution began universal molecular testing for newly diagnosed endometrial cancers in April 2023. In this study, we describe our institutional experience with this testing and specifically highlight POLEmut endometrial cancers. We aimed to compare the molecular and histopathologic profiles of POLEmut endometrial carcinomas with and without aggressive histopathologic features. A total of 198 endometrial cancer cases were analyzed by a targeted next-generation sequencing panel. We reviewed the results for all cases from April 2023 to December 2024 and for cases with pathogenic POLE exonuclease domain/proofreading mutation, collected relevant molecular, clinical data, immunohistochemical, and resection histopathology if available. Aggressive histopathology was defined as having at least one of the following: FIGO grade 3, ≥ stage pT1b, or lymphovascular invasion. Of 198 tested endometrial cancers, 40.9% (n=81) had no specific molecular profile, 33.8% (n=67) were p53 abnormal, 19.7% (n=39) were MMRd, and 5.6% (n=11) had POLE exonuclease domain/proofreading mutations associated with the ultramutated phenotype. At least one aggressive histopathologic feature was seen in 5/10 POLEmut cases that had resection specimens. TP53 variants were present in 5/11 POLE specimens and were often subclonal. Without sequencing, 4 POLEmut cases would have been misclassified as either p53 abnormal or MMRd. There was no significant difference in biomarker results, molecular variants, or clinical outcomes between cases with aggressive or nonaggressive histopathologic features.

Folate receptor 1 (FOLR1) has recently become a well-accepted therapeutic target in advanced-stage cancers. In this study, the prevalence of FOLR1 expression across all types of gynecologic tumors was investigated and correlated with selected clinicopathologic features. A total of 306 gynecologic tumors from 304 patients were evaluated for FOLR1 expression by immunohistochemistry (IHC). A positive FOLR1 is defined as ≥75% of viable tumor cells with moderate to strong membrane staining. Of 306 tumors, 31 (10.1%) had positive FOLR1 tests; a large majority of these FOLR1-positive tumors were HGSCs (64.5%), followed by uterine serous carcinoma, poorly differentiated/high-grade carcinoma, ovarian endometrioid carcinoma, ovarian mixed carcinoma, ovarian low-grade serous carcinoma, and serous borderline tumor with cribriform and micropapillary features. FOLR1 overexpression correlated with positive PD-L1 expression ( P =0.012), intact mismatch repair protein (MMR) expression ( P =0.024), and positive ER expression ( P =0.040). In endometrial tumors, positive FOLR1 expression was associated with poor histologic grade ( P =0.019), larger tumor size ( P =0.048), mutant p53 expression ( P <0.001), and lower PR expression ( P =0.015). Endometrial tumors with FOLR1 overexpression had a significantly higher rate of TP53 mutations ( P =0.013), while all endometrial tumors with PTEN alterations were negative for FOLR1 ( P =0.037). Overall, FOLR1 overexpression was associated with poor prognostic factors, such as advanced clinical stage, increased recurrence rate, higher pathologic T and N stage, poor histologic grade, larger tumor size, lymphovascular invasion, uterine serosa involvement, and shorter progression-free survival.

Endometrial cancer and uterine leiomyomas (fibroid) are common uterine pathologies that require early diagnosis to improve a patient's symptoms and increase the success rate of interventional procedures. In this work, we report on near-infrared spectroscopy (NIRS) spectral features of uterine cancer and fibroids from 69 surgical specimens obtained from 24 patients following hysterectomies. Normal uterus, cancer, and fibroid tissue were identified by NIR spectral contrast parameters based on the differences in spectrum morphology. Using the significant optical features and spectral principal components, a classification model was able to classify uterus tissue with a prediction accuracy higher than 70%, identifying cancer specimens with 70% sensitivity and 93% specificity, and fibroid samples with 86% sensitivity and 83% specificity. These results demonstrated NIRS mapping has promise as a complementary method for gynecologic imaging.

The uterus is central to the establishment, maintenance, and delivery of a healthy pregnancy. Biomechanics is an important contributor to pregnancy success, and alterations to normal uterine biomechanical functions can contribute to an array of obstetric pathologies. Few studies have characterized the passive mechanical properties of the gravid human uterus, and ethical limitations have largely prevented the investigation of mid-gestation periods. To address this key knowledge gap, this study seeks to characterize the structural, compositional, and time-dependent micro-mechanical properties of the nonhuman primate (NHP) uterine layers in nonpregnancy and at three time-points in pregnancy: early 2nd, early 3rd, and late 3rd trimesters. Distinct material and compositional properties were noted across the different tissue layers, with the nonpregnant endometrium and pregnant decidua being the least stiff, most viscous, least diffusible, and most hydrated layers of the NHP uterus. Pregnancy induced notable compositional and structural changes in the decidua and myometrium but had no effect on their micro-mechanical properties. Further comparison to published human data revealed marked similarities across species, with minor differences noted for the perimetrium and nonpregnant endometrium. This work provides insights into the material properties of the NHP uterus and demonstrates the validity of NHPs as a model for studying certain aspects of human uterine biomechanics.

Uterine fibroids are non-cancerous growths of the uterus that affect nearly 70%-80% of women in their lifetimes. Fibroids can cause severe pain, bleeding, and infertility. The main risk of recurrence is smaller fibroids, which are notoriously hard to detect, being missed during a surgical removal procedure, only to enlarge afterwards. In this work, hyperspectral imaging (HSI) datasets were acquired from samples from 10 patients after receiving a hysterectomy. Optical properties including absorption, scattering, and spectral morphology were extracted and fed into machine learning to classify regions as fibroid and myometrium. Top extracted optical features had significant contrast between fibroid and myometrium (p < 0.0001) and were used to train Random Forest (AUC: 0.9985 ± 0.001, Sensitivity: 0.9534 ± 0.019, Specificity: 0.9936 ± 0.009) and Logistic Regression (AUC: 0.9397 ± 0.013, Sensitivity: 0.8405 ± 0.023, Specificity: 0.8895 ± 0.032) with strong performance across testing splits. With HSI, there is contrast between fibroid and myometrium in the human uterus.

STUDY QUESTION:How does ovarian stimulation (OS), which is used to mature multiple oocytes for ART procedures, impact the principal cellular compartments and transcriptome of the human endometrium in the periovulatory and mid-secretory phases? SUMMARY ANSWER:During the mid-secretory window of implantation, OS alters the abundance of endometrial immune cells, whereas during the periovulatory period, OS substantially changes the endometrial transcriptome and impacts both endometrial glandular and immune cells. WHAT IS KNOWN ALREADY:Pregnancies conceived in an OS cycle are at risk of complications reflective of abnormal placentation and placental function. OS can alter endometrial gene expression and immune cell populations. How OS impacts the glandular, stromal, immune, and vascular compartments of the endometrium, in the periovulatory period as compared to the window of implantation, is unknown. STUDY DESIGN, SIZE, DURATION:This prospective cohort study carried out between 2020 and 2022 included 25 subjects undergoing OS and 25 subjects in natural menstrual cycles. Endometrial biopsies were performed in the proliferative, periovulatory, and mid-secretory phases. PARTICIPANTS/MATERIALS, SETTING, METHODS:Blood samples were processed to determine serum estradiol and progesterone levels. Both the endometrial transcriptome and the principal cellular compartments of the endometrium, including glands, stroma, immune, and vasculature, were evaluated by examining endometrial dating, differential gene expression, protein expression, cell populations, and the three-dimensional structure in endometrial tissue. Mann-Whitney U tests, unpaired t-tests or one-way ANOVA and pairwise multiple comparison tests were used to statistically evaluate differences. MAIN RESULTS AND THE ROLE OF CHANCE:In the periovulatory period, OS induced high levels of differential gene expression, glandular-stromal dyssynchrony, and an increase in both glandular epithelial volume and the frequency of endometrial monocytes/macrophages. In the window of implantation during the mid-secretory phase, OS induced changes in endometrial immune cells, with a greater frequency of B cells and a lower frequency of CD4 effector T cells. LARGE SCALE DATA:The data underlying this article have been uploaded to the Genome Expression Omnibus/National Center for Biotechnology Information with accession number GSE220044. LIMITATIONS, REASONS FOR CAUTION:A limited number of subjects were included in this study, although the subjects within each group, natural cycle or OS, were homogenous in their clinical characteristics. The number of subjects utilized was sufficient to identify significant differences; however, with a larger number of subjects and additional power, we may detect additional differences. Another limitation of the study is that proliferative phase biopsies were collected in natural cycles, but not in OS cycles. Given that the OS cycle subjects did not have known endometrial factor infertility, and the comparisons involved subjects who had a similar and robust response to stimulation, the findings are generalizable to women with a normal response to OS. WIDER IMPLICATIONS OF THE FINDINGS:OS substantially altered the periovulatory phase endometrium, with fewer transcriptomic and cell type-specific changes in the mid-secretory phase. Our findings show that after OS, the endometrial microenvironment in the window of implantation possesses many more similarities to that of a natural cycle than does the periovulatory endometrium. Further investigation of the immune compartment and the functional significance of this cellular compartment under OS conditions is warranted. STUDY FUNDING/COMPETING INTERESTS:Research reported in this publication was supported by the National Institute of Allergy and Infectious Diseases (R01AI148695 to A.M.B. and N.C.D.), Eunice Kennedy Shriver National Institute of Child Health and Human Development (R01HD109152 to R.A.), and the March of Dimes (5-FY20-209 to R.A.). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health or March of Dimes. All authors declare no conflict of interest.

BACKGROUND Angiosarcoma is a rare malignant tumor. Owing to the lack of specific clinical manifestations of this disease, it is difficult to achieve early diagnosis and start early treatment. CASE SUMMARY A 78-year-old male patient was admitted to the hospital because of a bump on his head that did not heal for 4 mo. The patient was diagnosed with a refractory head wound. The patient underwent neoplasm resection and skin grafting surgery in the Plastic Surgery. The neoplasm was sent for pathological examination during the operation. The final pathological results were confirmed scalp angiosarcoma. CONCLUSION Our research suggests that pathological examination should be performed for refractory ulcers of the scalp, and physical factor therapy should be used with caution before the diagnosis is clear.

Ovarian stimulation (OS) exposes the endometrium to supraphysiologic levels of estradiol (E2) and an early rise in progesterone (P4). We examined the glandular, stromal, vascular and immune compartments of the endometrium in order to better understand the mechanisms underlying OS-mediated changes. We hypothesized that OS-mediated changes would be apparent as early as the periovulatory (PO) endometrium and that these changes would persist into the mid-secretory (MSE) phase leading to endometrial aberrations during the window of receptivity.

Perivascular epithelioid cell tumors (PEComas), rare mesenchymal tumors with myomelanocytic differentiation, can be a diagnostic challenge, often requiring a panel of immunohistochemical markers. Preferentially expressed antigen in melanoma (PRAME) is a relatively new antigen with utility in diagnosing melanomas. This study aimed to survey PRAME expression patterns in the PEComa family of tumors and morphologic mimics. Twenty cases of PEComas and 27 non-PEComas (10 leiomyosarcomas, 3 smooth muscle tumors of uncertain malignant potential [STUMPs], 11 leiomyomas, 1 uterine inflammatory myofibroblastic tumor [IMT], and 2 low-grade endometrial stromal sarcomas [LGESSs]) were stained with PRAME and compared to previously performed HMB45 and Melan-A stains, when available. Tumors showing no or barely perceptible PRAME staining at 10× were considered negative. Tumors were considered positive if there was full nuclear staining evident at 10× in at least one 10× field. Diffuse staining was defined as positivity in at least 80% of tumor nuclei. Overall, PRAME was expressed in 70% of PEComas, with diffuse positivity in 60%. However, PRAME was not specific for PEComas, with immunopositivity in the majority (70%) of uterine leiomyosarcoma cases, though negative in STUMP, leiomyoma, IMT, and LGESS cases. PRAME sensitivity was 70% and specificity was 74%, while HMB45 was more sensitive (90%) and specific (100%), but only 15% of PEComas showed diffuse staining. Melan-A staining was less common than HMB45 or PRAME, with only 18.8% sensitivity but 100% specificity. Among gynecologic PEComas, PRAME was expressed in 75% overall and enriched among malignant cases (85.7% positive). As part of an immunohistochemical panel, PRAME could be useful in the workup of PEComa cases. In the future, PRAME-specific immunotherapies may be beneficial in treating patients with malignant PEComas.

e17559 Background: High grade serous ovarian cancer (HGSC) is the most common and fatal epithelial ovarian cancer and is often diagnosed in late stages. DNA methylation has emerged as a potential biomarker for the early detection of cancer, including ovarian cancer. Studies examining DNA methylation in ovarian tumor tissue have used adjacent non-tumor tissues or tissues from unaffected women as the control; however, few have used paired tissue and research is sparse on how results may vary by the control non-tumor tissue type. Therefore, we examined DNA methylation signatures in HGSC tumor tissue using different control non-tumor tissue types. Methods: We examined DNA methylation signatures using the Illumina Infinium MethylationEPIC BeadChip (850k) in formalin-fixed paraffin-embedded tissue. In women with HGSC, we compared DNA methylation patterns between paired adjacent non-tumor ovary (n = 74) and fallopian tube (n = 80) tissues. We compared DNA methylation patterns in these adjacent non-tumor tissues with non-tumor ovary (n = 8) and fallopian tube (n = 8) tissues from unaffected women without ovarian cancer carrying a pathogenic variant in BRCA1/2 ( BRCA1/2+). Lastly, we compared the overlap in differentially methylated CpGs identified in HGSC tumor tissue compared to paired adjacent non-tumor ovary (n = 50) and fallopian tube (n = 52) tissues. We processed the methylation data and used principal components analysis (PCA) and the adjusted Rand Index to compare the non-tumor tissue type methylation patterns. We used a paired t-test between individual-matched tumor and adjacent non-tumor tissue for each CpG site and then applied Bonferroni’s method to adjust the obtained p-values. Results: PCA comparing methylation patterns between paired adjacent non-tumor ovary and fallopian tube tissues from women with HGSC showed an adjusted Rand Index of 0.78, revealing separate clusters that cannot be combined. PCA comparing methylation patterns from the adjacent non-tumor tissues from women with HGSC and the non-tumor tissues from unaffected BRCA1/2+ women showed an adjusted Rand Index of -0.10 and 0.07 for the ovary and fallopian tube tissues, respectively, revealing overlapping clusters that can be combined. Lastly, comparison of the top differentially methylated CpGs identified in HGSC tumor tissue when using paired adjacent non-tumor ovary versus fallopian tube tissues as the control showed minimal overlap (6.8% and 4.0% for hypermethylated and hypomethylated CpGs, respectively). Conclusions: These results suggest that paired adjacent non-tumor ovary and fallopian tube tissues from women with HGSC have different DNA methylation patterns and result in different methylation signatures identified in HGSC tumor tissue. When comparing results across studies, including for validation, the type of non-tumor tissue control must be considered.

Mesonephric-like adenocarcinomas (MLA) are rare neoplasms arising in the cervix, endometrium, and ovary. In contrast to mesonephric carcinomas (MC), mesonephric-like adenocarcinomas are not associated with mesonephric remnants. Both entities have a similar appearance with regards to varying histomorphology patterns, including glandular, tubular, spindled, solid, and papillary, and have a specific immunophenotype and molecular features. We present a case of a 54-year-old HPV-negative woman with a Pap test that exhibits high-grade malignancy. The cell block displayed malignant cells with positive stains for PAX8, GATA3, and TTF1 by immunohistochemistry. The diagnosis of adenocarcinoma with mesonephric like features was rendered. MLA can be challenging on the small specimens and often misinterpreted as other endometrial neoplasms. Furthermore, the accurate diagnosis carries a well-described risk of aggressive clinical behavior.

Spontaneous preterm birth (sPTB), a major cause of infant morbidity and mortality, must involve premature cervical softening/dilation for a preterm vaginal delivery to occur. Yet, the mechanism behind premature cervical softening/dilation in humans remains unclear. We previously reported the non-pregnant human cervix contains considerably more cervical smooth muscle cells (CSMC) than historically appreciated and the CSMC organization resembles a sphincter. We hypothesize that premature cervical dilation leading to sPTB may be due to (1) an inherent CSMC contractility defect resulting in sphincter failure and/or (2) altered cervical extracellular matrix (ECM) rigidity which influences CSMC contractility. To test these hypotheses, we utilized immunohistochemistry to confirm this CSMC phenotype persists in the human pregnant cervix and then assessed in vitro arrays of contractility (F:G actin ratios, PDMS pillar arrays) using primary CSMC from pregnant women with and without premature cervical failure (PCF). We show that CSMC from pregnant women with PCF do not have an inherent CSMC contractility defect but that CSMC exhibit decreased contractility when exposed to soft ECM. Given this finding, we used UPLC-ESI-MS/MS to evaluate collagen cross-link profiles in the cervical tissue from non-pregnant women with and without PCF and found that women with PCF have decreased collagen cross-link maturity ratios, which correlates to softer cervical tissue. These findings suggest having soft cervical ECM may lead to decreased CSMC contractile tone and a predisposition to sphincter laxity that contributes to sPTB. Further studies are needed to explore the interaction between cervical ECM properties and CSMC cellular behavior when investigating the pathophysiology of sPTB.

Aims The wide variety of affected organ systems associated with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection highlights the need for tissue-specific evaluation. We compared placentas from SARS-CoV-2-positive and SARS-CoV-2-negative women in our hospital in New York City, which became the epicenter of the coronavirus disease 2019 pandemic in March 2020. To date, some limited studies have been published on placentas from SARS-CoV-2-positive women. The aim of our study, in addition to describing histomorphology, was to utilizein-situhybridization (ISH) for the S-gene encoding the spike protein and immunohistochemistry (IHC) with the monoclonal SARS-CoV-2 spike antibody 1A9 for placental evaluation. Methods and results In this study, 51 singleton, third-trimester placentas from SARS-CoV-2-positive women and 25 singleton, third-trimester placentas from SARS-CoV-2-negative women were examined histomorphologically according to the Amsterdam Criteria and with ISH and/or IHC. The corresponding clinical findings and neonatal outcomes also were recorded. Although no specific histomorphologic changes related to SARS-CoV-2 were noted in the placentas, evidence of maternal-fetal vascular malperfusion was identified, with placentas from SARS-CoV-2-positive women being significantly more likely to show villous agglutination (P = 0.003) and subchorionic thrombi (P = 0.026) than placentas from SARS-CoV-2-negative women. No evidence of direct viral involvement was identified with ISH and IHC. Conclusions In this study, third-trimester placentas from SARS-CoV-2-positive women were more likely to show evidence of maternal-fetal vascular malperfusion; however, ISH and IHC provided no evidence of direct viral involvement or vertical transmission.

Sex cord tumor with annular tubules (SCTAT) is a rare ovarian tumor with characteristic microscopic morphologic features. Diagnosis most often is based on examination of tissue specimens. Cytologic features of this tumor rarely have been described in the English literature. Herein, we report cytologic findings of cyst aspiration fluid in two cases of SCTAT, with cyto-histologic correlation.