Expression and Prognostic Significance of LAG-3, TIGIT, VISTA, and IDO1 in Endometrial Serous Carcinoma.

Endometrial serous carcinoma (ESC) is an uncommon, aggressive type of endometrial cancer. While immune checkpoint blockade has emerged as a promising treatment option for endometrial carcinomas, research on the expression of immune checkpoints that could serve as prospective immunotherapy targets in ESC is limited. We examined the prevalence and prognostic value of lymphocyte-activation gene 3 (LAG-3), T-cell immunoglobulin and ITIM domain (TIGIT), V-domain immunoglobulin (Ig) suppressor of T-cell activation (VISTA), and indoleamine 2,3-dioxygenase 1 in 94 cases of ESC and correlated their expression with CD8+ and FOXP3+ tumor-infiltrating lymphocytes (TILs). We observed a positive correlation among LAG-3, TIGIT, and VISTA expressed on immune cells, and among these markers and CD8+ and FOXP3+ TIL densities. In Kaplan-Meier survival analysis, tumors with high levels of LAG-3 and TIGIT expression had better progression-free survival (PFS) and overall survival (OS) than those with lower levels of expression (LAG-3: PFS, P = .03, OS, P = .04; TIGIT: PFS, P = .01, OS, P = .009). In multivariate analysis, only high TIGIT expression was of independent prognostic value for better OS. VISTA expression in immune or tumor cells, and indoleamine 2,3-dioxygenase 1 expression in tumor cells, did not show a significant association with survival. Our data indicate that LAG-3, TIGIT, and VISTA immune checkpoints have roles in the microenvironment of ESC, and their expression patterns highlight the complex interactions among the different components of this system. High levels of these markers, together with high CD8+ TIL, suggest the potential immunogenicity of a subset of these tumors. Further studies are needed to elucidate the roles of various immune components in the ESC microenvironment and their association with intrinsic tumor properties.

Predicting Progestin Therapy Response With PTEN, PAX2, and ß-Catenin in Patients With Endometrioid Precancer.

This study investigates the predictive value of biomarkers PTEN, PAX2, and ß-catenin for therapeutic outcomes in patients with atypical endometrial hyperplasia or endometrioid intraepithelial neoplasia undergoing progestin therapy. In a retrospective study of 128 patients, we analyzed a total of 351 endometrial biopsy samples and categorized outcomes into responders (absence of residual disease) and nonresponders (presence of residual disease). We found aberrant biomarker expression in pretreatment cases: 48% for PTEN, 65% for PAX2, and 36% for ß-catenin. Approximately 77.3% of patients responded to progestin treatment, with nonresponders showing significantly higher initial PTEN loss (75.86% vs 39.79%, P < 0.001). Nonresponders also demonstrated significant PTEN loss (53.33% vs 20.55%, P < 0.001), PAX2 loss (57.33% vs 41.22%, P < 0.05), and ß-catenin nuclear staining (53.45% vs 27.91%, P < 0.01) in follow-up samples. In addition, nonresponders exhibited lower recovery of intact PTEN and PAX2, along with higher ß-catenin aberrancy in cases initially showing normal ß-catenin levels. We conclude that persistent aberrant PTEN and PAX2 expression, coupled with emerging aberrant ß-catenin in follow-ups, indicates a greater likelihood of treatment failure. Conversely, the absence of these aberrations suggests successful progestin therapy. Our findings highlight the utility of this 3-marker panel in assessing residual disease status and predicting progestin treatment outcomes, thus offering critical insights for patient management.

Somatic and autonomic nerve density and distribution within the clitoris: an immunohistochemical study in adult female cadavers.

INTRODUCTION AND HYPOTHESIS: Knowledge of clitoral neuroanatomy is critical to vulvar surgery. We sought to characterize the density and distribution of autonomic and somatic nerves supplying the clitoris. METHODS: Pelvic tissue harvested from female cadavers was sectioned axially at three anatomic levels: the proximal aspect of the clitoral body (CB), the distal CB, and the glans. The CB, glans, and the surrounding connective tissue (dorsal, lateral, and ventral) were outlined microscopically. An area containing large nerve bundles dorsal to the CB, referred to as the dorsal nerve subregion, was analyzed separately. Double-immunofluorescent staining for beta III tubulin (ßIIIT), a global axonal marker, and myelin basic protein (MBP), a myelinated nerve marker, was performed. Threshold-based automatic image-segmentation distinguished stained areas. Autonomic and somatic density were calculated as percentage of tissue stained with ßIIIT alone, and ßIIIT and MBP respectively. Comparisons were made using nonparametric Friedman tests. RESULTS: Seven cadavers, aged 22-81, were examined. Somatic (mean 4.42%, SD ± 1.97) and autonomic (2.14% ± 2.42) nerve density was highest in the dorsal nerve subregion and dorsal region at the distal CB level. Compared with the CB, somatic density was higher in proximal (0.05% ± 0.03 vs 1.27% ± 0.69, p = 0.03) and distal (0.29% ± 0.25 vs 1.09% ± 0.41, p = 0.05) dorsal regions. Somatic density was greater in the glans than in the surrounding lateral (0.78% ± 0.47 vs 0.43% ± 0.23, p = 0.03) and ventral (0.78% ± 0.47 vs 0.52% ± 0.2, p = 0.03) regions. Autonomic density was greater than somatic in all areas, except for the dorsal nerve subregion. CONCLUSIONS: Somatic and autonomic nerve density were greatest in a well-defined region dorsal to the CB. Surgical preservation of this region is critical for maintaining nerve supply to the clitoris.

Anatomic relationships of the clitoral body, bulbs of the vestibule, and urethra.

BACKGROUND: Although recent studies have enhanced our understanding of the anatomy of the clitoris and its somatic innervation, less emphasis has been placed on the anatomic relationships of the clitoris to its surrounding structures. OBJECTIVE: This study aimed to further characterize the gross and histologic relationships of the clitoris, vestibular bulbs, and urethra. STUDY DESIGN: Detailed dissections were performed in 30 unembalmed female cadavers. In 23 specimens, gross dissections were performed, and relationships of the clitoris, vestibular bulbs, and urethra were annotated. Histologic evaluation was performed in 7 specimens, in which tissues were harvested within 24 hours from death. Descriptive statistics were used for data analyses. RESULTS: The clitoral body consisted of 2 components, the proximal body and the distal body. The distal body was oriented ≤90° from the proximal body, forming an outer and inner angle at the inflection point. A "septumlike" arrangement of fibroconnective and vascular tissues was noted between the inner angle of the clitoral body and the urethra. Neurovascular bundles coursed laterally along the clitoral body and the surfaces of the crura and vestibular bulbs. The vestibular bulbs approached each other over the ventral surface of the urethra, at the commissure of the vestibular bulbs. Each bulb was separated by fibrous tissue and did not merge along the midline. The vestibular bulbs approximated the clitoral body, but the erectile tissue of the vestibular bulbs was separated from the corpora cavernosa of the clitoral body by the tunica albuginea. The erectile tissue of the vestibular bulbs abutted the ventrolateral walls of the urethra but was separated from the urethral mucosa by an indiscrete layer of erectilelike tissue with dense stroma. CONCLUSION: This study provided gross and histological confirmation of the relationships of the clitoris, vestibular bulbs, and urethra. Detailed knowledge of the anatomy of the clitoris is crucial for reducing surgical complications associated with periclitoral and distal urethral procedures, which may adversely affect sexual arousal and sexual function.

Somatic and autonomic nerve density of the urethra, periurethral tissue, and anterior vaginal wall: an immunohistochemical study in adult female cadavers.

INTRODUCTION AND HYPOTHESIS: Retropubic procedures may disrupt nerves supplying the pelvic viscera; however, knowledge of pelvic neuroanatomy is limited. We sought to characterize somatic and autonomic nerve density within the urethra, periurethral tissue, and anterior vagina. METHODS: Axial sections were obtained from pelvic tissue harvested from female cadavers ≤24 h from death at three anatomical levels: the midurethra, proximal urethra, and upper trigone. Periurethral/perivesical tissue was divided into medial and lateral sections, and the anterior vagina into middle, medial, and lateral sections. Double immunofluorescent staining for beta III tubulin (ßIIIT), a global axonal marker, and myelin basic protein (MBP), a myelinated nerve marker, was performed. Threshold-based automatic image segmentation distinguished stained areas. Autonomic and somatic density were calculated as percentage of tissue stained with ßIIIT alone, and with ßIIIT and MBP respectively. Statistical comparisons were made using nonparametric Friedman tests. RESULTS: Six cadavers, aged 22-73, were examined. Overall, autonomic nerve density was highest at the midurethral level in the lateral and middle anterior vagina. Somatic density was highest in the external urethral sphincter (midurethra mean 0.15%, SD ±0.11; proximal urethra 0.19%, SD ±0.19). Comparison of annotated sections revealed significant differences in autonomic density among the lateral, medial, and middle vagina at the midurethra level (0.71%, SD ±0.48 vs 0.60%, SD ±0.48 vs 0.70%, SD ±0.63, p=0.03). Autonomic density was greater than somatic density in all sections. CONCLUSIONS: Autonomic and somatic nerves are diffusely distributed throughout the periurethral tissue and anterior vagina, with few significant differences in nerve density among sections analyzed. Minimizing tissue disruption near urethral skeletal muscle critical for urinary continence may prevent adverse postoperative urinary symptoms.

Prevalence and prognostic significance of PD-L1, TIM-3 and B7-H3 expression in endometrial serous carcinoma.

Endometrial serous carcinoma (ESC) is an aggressive type of endometrial carcinoma with a poor prognosis. Immune checkpoint blockade has evolved as a novel treatment option for endometrial cancers; however, data on expression of immune checkpoints that may be potential targets for immunotherapy in ESC are limited. We analyzed the prevalence and prognostic significance of PD-L1, TIM-3 and B7-H3 immune checkpoints in 99 ESC and evaluated their correlation with CD8 + tumor infiltrating lymphocytes. Applying the tumor proportion score (TPS) with a cutoff of 1%, PD-L1, TIM-3 and B7-H3 expression was present in 17%, 10% and 93% of cases, respectively. Applying the combined positive score (CPS) with a cutoff of 1, PD-L1, TIM-3 and B7-H3 expression was present in 63%, 67% and 94% of cases, respectively. Expression of these markers was largely independent of one another. PD-L1 correlated with higher CD8 + T-cell density when evaluated by either TPS (p = 0.02) or CPS (p < 0.0001). TIM-3 correlated with CD8 + T-cell density when evaluated by CPS (p < 0.0001). PD-L1 positivity was associated with improved overall survival (p = 0.038) when applying CPS. No association between PD-L1 expression and survival was found using TPS, and there was no association between TIM-3 or B7-H3 positivity and survival by either TPS or CPS. Using TPS, PD-L1 correlated with a higher tumor stage but not with survival, whereas the converse was true when PD-L1 was evaluated by CPS, suggesting that PD-L1 expression in immune cells correlates with prognosis and is independent of tumor stage. In conclusion, PD-L1, TIM-3 and B7-H3 may be potential therapeutic targets in selected patients with ESC. Further investigation of their roles as predictive biomarkers is needed.

Rare Complete Hydatidiform Mole With p57 Expression in Villous Mesenchyme: Case Report and Review of Discordant p57 Expression in Hydatidiform Moles.

Complete hydatidiform mole (CHM) is a premalignant proliferative disease of the placenta characterized by misexpression of imprinted gene products, most notably p57. The majority of CHM exhibit immunohistochemical absence of p57 protein in villous mesenchyme (VM) and cytotrophoblast (CT) and are thus p57 VM/CT concordant. However, some gestations show loss of p57 in only VM or CT, either in all chorionic villi or a subset thereof (VM/CT discordant). Here, we present a rare case of a p57 VM/CT-discordant CHM with diffuse retention of p57 expression in VM but complete absence in CT. Histologically, the case exhibited typical features of CHM including trophoblast hyperplasia and severe nuclear atypia, but was unusual in the presence of gestational membranes identified ultrasonographically and histologically. Ploidy determination by FISH and genotyping by short tandem repeat analyses showed that this was a diploid gestation with variable allelic ratios and with an androgenetic lineage, similar to previously reported p57 VM/CT-discordant cases.

Low grade endometrial endometrioid adenocarcinoma: A review and update with emphasis on morphologic variants, mimics, immunohistochemical and molecular features.

Endometrioid adenocarcinoma (ECa) may feature a number of morphologic variations that can pose diagnostic challenge. The purpose of this review and update is to examine the spectrum of morphologic variants and mimics of low-grade (FIGO grades 1 and 2) ECa, with a focus on histologic, immunohistochemical, and molecular features that may inform diagnosis and treatment. In addition to ECa of usual type, variants with unique cytologic and/or architectural features presented include the following: 1) ECa with mucinous differentiation of conventional (Müllerian) type; 2) ECa with squamous differentiation; 3) ECa with morular metaplasia; 4) ECa with patterns resembling cervical transformation zone tissue and/or microglandular hyperplasia; 5) ECa with cytoplasmic clearing; 6) ECa with papillation, including villoglandular variant of ECa, ECa with small nonvillous papillae, and ECa with a "low-grade serous"-like component or surface changes mimicking ovarian serous borderline tumor; 7) corded and hyalinized variant of ECa; 8) ECa with spindled epithelial cells; 9) ECa with sex cord-like pattern; and 10) ECa with other unusual cytologic and associated features. For each variant, relevant differential diagnoses and diagnostic strategies are discussed. The most clinically significant distinctions come into play in the differential diagnosis between low-grade ECa and one of its high-grade mimics. In this setting, the most fundamental tool in the pathologist's diagnostic arsenal is recognition of the low-grade cytologic features typical of low-grade ECa. Circumspect evaluation of cytologic features, complemented by an awareness of the morphologic spectrum, an appropriate battery of immunohistochemical stains when needed, and mindfulness of the clinical scenario, should guide the pathologist to the correct histotype in even challenging cases.

Navigating through perplex morphologic changes after exogenous hormone usage.

Clinical application of exogenous hormone as a method of contraception and/or treatment of various gynecologic disorders is exceedingly common. Unfortunately, the concurrent use of these agents also complicates the interpretation of pathology specimens. Various studies have shown that morphologic changes induced by hormonal therapies are present in both non-neoplastic and neoplastic tissues within the women's reproductive tract. It is important to understand the exogenous hormone induced morphologic changes, as it helps the pathologists make the accurate diagnosis, and in turn, guide clinicians to make optimal clinical decisions. In this review, we summarize the morphologic changes in both neoplastic and non-neoplastic endometrial, cervical, and myometrial surgical specimens after hormonal therapies, particularly after progestin treatment. In the endometrium, particularly in the scenario of progestin-treated atypical endometrial hyperplasia/endometrioid intraepithelial neoplasia (AEH/EIN), there is notoriously poor interobserver agreement and difficulty in assessing for the residual disease. We summarize current literature and propose our recommended approach in assessing these challenging endometrial biopsies, including a diagnostic algorism, the use of PAX-2, PTEN, beta-catenin immunohistochemistry panel, as well as consistency in diagnostic wording of the report. In the cervix, progestin makes dysplastic lesions appear metaplastic, thus high-grade squamous dysplastic lesions may be easily missed. Within the myometrium, lesions such as adenomyosis may show various degree of decidualization, while smooth muscle neoplasms may show apoplectic changes, and stromal lesions including endometrial stromal sarcoma may show more eosinophilic cytoplasm. All such changes may pose more or less diagnostic challenges in our daily practice. However, most are readily recognizable when we understand particular hormone related scenarios.

Anatomy, histology, and nerve density of clitoris and associated structures: clinical applications to vulvar surgery.

BACKGROUND: A precise understanding of structures comprising the female external genitalia is essential in obstetric and gynecologic practice. OBJECTIVE: To further characterize the anatomy, histology, and nerve density of the clitoris and associated structures, and to provide clinical correlations to vulvar surgery. MATERIALS AND METHODS: Unembalmed female cadavers were examined. The length and width of the body, glans, and crura of the clitoris were measured. Distances from the glans to the urethra and from the dorsal surface of the clitoral body to the mid pubic arch were recorded. The path of the dorsal nerve of the clitoris was examined, and the nerve width was measured as it emerged from the lateral surface of crura and at the distal clitoral body. Distances from where the dorsal nerve emerged from the perineal membrane to the posterior surface of the membrane and to mid pubic arch were measured. Connective tissue layers associated with the clitoris were examined. Tissue was harvested from additional unembalmed cadavers, and nerve density of the labia minora, glans, and clitoral body were analyzed. Histological examination was performed on vulvar structures to clarify tissue composition. Descriptive statistics were used for data analyses. RESULTS: A total of 27 cadavers (aged 48-96 years) were examined, 22 grossly and 5 histologically. The median length and width of clitoral body were 29 mm (range, 13-59 mm) and 9 mm (range, 5-14 mm), respectively. The glans was 8 mm (range, 5-12 mm) long and 4 mm (range, 3-10 mm) wide. The length of the crura was 50 mm (range, 25-68 mm), and the width at the anterior portion was 9 mm (range, 2-13 mm). The closest distance from the glans to the urethra was 25 mm (range, 14-37 mm) and from the clitoral body to the mid pubic arch was 29 mm (range, 14-46 mm). The widths of the dorsal nerve at the lateral crura and at the distal clitoral body were 3 mm (range, 2-4 mm) and 1 mm (range, 1-2 mm), respectively. The distance from the dorsal nerve as it emerged from the perineal membrane to the mid pubic arch was 34 mm (range, 20-48 mm) and to the posterior surface of the membrane was 20 mm (range, 8-31 mm). The dorsal nerve and artery of the clitoris coursed adjacent to the medial surface of the inferior pubic ramus surrounded by a dense fibrous capsule adherent to the periosteum. The nerve and artery then coursed deep to dense connective tissue layers, which were contiguous with the suspensory ligament and fascia of the clitoris. Histologic examination revealed the presence of erectile tissue in the clitoral body, crura, and vestibular bulbs, but such tissue was absent in the glans and labia minora. Nerve density analysis revealed statistically significant greater density in the dorsal compared with ventral half of the clitoral body. Although not statistically significant, there was increased nerve density in the distal compared to the proximal half of the labia minora. CONCLUSION: Precise knowledge of clitoral anatomy and associated neurovascular structures is essential to safely complete partial vulvectomies, clitoral and vulvar reconstructive procedures, anti-incontinence surgeries, and repair of obstetric lacerations. Understanding the range of anatomic variations and awareness of the areas of increased nerve density is important during counseling and surgical planning. Although the dorsal nerve of the clitoris courses deep to dense connective tissue layers, inadvertent injury may occur in the setting of deep dissection or suture placement. The dorsal nerve seems most vulnerable with surgical entry or lacerations that extend from the midline of the prepuce to the inferior pubic rami.

Gross and histologic relationships of the retropubic urethra to lateral pelvic sidewall and anterior vaginal wall in female cadavers: clinical applications to retropubic surgery.

BACKGROUND: Knowledge of the retropubic space anatomy is essential for safe entry and surgical applications within this space. OBJECTIVE: The objectives of this study were to examine the gross and histologic anatomy of the retropubic urethra, paraurethral tissue, and urethrovaginal space and to correlate findings to retropubic procedures. STUDY DESIGN: Anatomic relationships of the retropubic urethra were examined grossly in unembalmed female cadavers. Measured distances included: lateral urethral wall to arcus tendineus fascia pelvis at the level of urethrovesical junction and at 1 cm distal. Other measurements included retropubic urethral length and distances from internal urethral opening to each ureteric orifice. Microscopic examination was performed at the same levels examined grossly in separate nulliparous specimens. Descriptive statistics were used for data analyses. RESULTS: In all, 25 cadavers were examined grossly. Median distance from lateral urethral wall to arcus tendineus fascia pelvis at the level of urethrovesical junction was 25 mm (range, 13-38 mm). At 1 cm distal, the median distance from aforementioned structures was 14 mm (10-26 mm). Median length of the retropubic urethra was 23 mm (range 15-30 mm). Four nulliparous specimens, ages 12 weeks, and 34, 47, and 52 years, were examined histologically. No histologic evidence of a discrete fascial layer between bladder/urethra and anterior vagina was noted at any level examined. Tissue between the urethra and the pelvic sidewall skeletal muscle was composed of dense fibrous tissue, smooth muscle bundles, scant adipose tissue, blood vessels, and nerves. The smooth muscle fibers of the vaginal muscularis interdigitated with skeletal muscle fibers in the pelvic sidewall at both levels examined. No histologic evidence of "pubourethral ligaments" within the paraurethral tissue was noticed. CONCLUSION: A 2-cm "zone of safety" exists between the urethra and arcus tendineus fascia pelvis at the urethrovesical junction level. Suture or graft placement within this region should minimize injury to the urethra, pelvic sidewall muscles, and bladder. Knowledge that the shortest length of retropubic urethra was 1.5 cm and shortest urethra to arcus tendineus fascia pelvis distance was 1 cm highlights the importance of maintaining dissection and trocar entry site close to pubic bone to avoid bladder and/or urethral injury. Histologic analysis of paraurethral tissue supports the nonexistence of pubourethral ligaments.

Anatomic relationships of the pelvic autonomic nervous system in female cadavers: clinical applications to pelvic surgery.

BACKGROUND: The integrity of the pelvic autonomic nervous system is essential for proper bowel, bladder, and sexual function. OBJECTIVE: The purpose of this study was to characterize the anatomic path of the pelvic autonomic system and to examine relationships to clinically useful landmarks. STUDY DESIGN: Detailed dissections were performed in 17 female cadavers. Relationships of the superior hypogastric plexus to aortic bifurcation and midpoint of sacral promontory were examined; the length and width of plexus was documented. Path and width of right and left hypogastric nerves were recorded. The origin and course of the pelvic splanchnic nerves were documented. Individual nerve tissue that contributed to the inferior hypogastric plexus was noted. Relative position of nerves to arteries, viscera, and ligaments was documented. In a subset of specimens, biopsy specimens were obtained to confirm gross findings by histologic analysis. Descriptive statistics were used for data analyses and reporting. RESULTS: In all specimens, the superior hypogastric plexus was embedded in a connective tissue sheet within the presacral space, just below the peritoneum. In 14 of 17 specimens (82.4%), the plexus formed a median distance of 21.3 mm (range, 9-40 mm) below aortic bifurcation; in the remaining specimens, it formed a median distance of 25.3 mm (range, 20.5-30 mm) above bifurcation. In 58.8% of specimens, the superior hypogastric plexus was positioned to the left of midline. The median length and width of the plexus was 39.5 (range, 11.5-68) mm and 9 (range, 2.5-15) mm, respectively. A right and left hypogastric nerve was identified in all specimens and formed a median distance of 23 mm (range, 5-32 mm) below the promontory. The median width of the hypogastric nerve was 3.5 mm (range, 3-4.5 mm) on the right and 3.5 mm (range, 2-6.5 mm) on the left. The median distance from midportion of uterosacral ligament to the closest nerve branch was 0.5 mm (range, 0-4.5 mm) on right and 0 mm (range, 0-27.5 mm) on left. In all specimens, the inferior hypogastric plexus was formed by contributions from the hypogastric nerves and branches from S3 and S4. In 47.1% of hemipelvises, S2 branches contributed to the plexus. The sacral sympathetic trunk contributed to the plexus in 16 of 34 hemipelvises where this structure was identified. The inferior hypogastric plexus formed 1-3 cm lateral to the rectum and upper third of the vagina. From this plexus, 1-3 discrete branches coursed deep to the ureter toward the bladder. A uterine branch that coursed superficial to the ureter followed the ascending branch of the uterine artery. An S4 branch was found directly attaching to lateral walls of the rectum in 53% of specimens. Pelvic splanchnic nerves merged into the inferior hypogastric plexus on the lower and medial surface of the coccygeus muscle. Histologic analysis confirmed neural tissue in all tissues that were sampled. CONCLUSION: Anatomic variability and inability to visualize the small caliber fibers that comprise the inferior hypogastric plexus grossly likely underlines the reasons that some postoperative visceral and sexual dysfunction occur in spite of careful dissection and adequate surgical technique. These findings highlight the importance of a discussion with patients about the risks that are associated with interrupting autonomic fibers during the preoperative consent.

Utility of a PAX2, PTEN, and ß-catenin Panel in the Diagnosis of Atypical Hyperplasia/Endometrioid Intraepithelial Neoplasia in Endometrial Polyps.

The diagnosis of atypical hyperplasia/endometrioid intraepithelial neoplasm (AH/EIN) within endometrial polyps (EMPs) often poses a diagnostic conundrum. Our previous studies demonstrated that a panel of immunohistochemical (IHC) markers consisting of PAX2, PTEN, and ß-catenin can be effectively utilized for the identification of AH/EIN. A total of 105 AH/EIN within EMP were analyzed using the 3-marker panel. We also evaluated these cases for the presence of morules. Benign EMP (n=90) and AH/EIN unassociated with polyp (n=111) served as controls. Aberrant expression of PAX2, PTEN, or ß-catenin was observed in AH/EIN in EMP in 64.8%, 39.0%, and 61.9% of cases, respectively. At least 1 IHC marker was abnormal in 92.4% of cases. Overall, 60% of AH/EIN in EMP demonstrated abnormal results for≥2 IHC markers. The prevalence of PAX2 aberrancy was significantly lower in AH/EIN in EMP than in nonpolyp AH/EIN (64.8% vs. 81.1%, P =0.007), but higher than in benign EMP (64.8% vs. 14.4%, P <0.00001). The prevalence of ß-catenin aberrancy was significantly higher in AH/EIN in EMP than in nonpolyp AH/EIN (61.9% vs. 47.7%, P =0.037). All control benign EMP demonstrated normal expression of PTEN and ß-catenin. Morules were present in 38.1% of AH/EIN in EMP versus 24.3% in nonpolyp AH/EIN, and absent in benign EMP. A strong positive association was found between ß-catenin and morules (Φ=0.64). Overall, 90% cases of atypical polypoid adenomyoma (n=6) and mucinous papillary proliferation (n=4) showed IHC marker aberrancy. In conclusion, the 3-marker IHC panel (PAX2, PTEN, and ß-catenin) is (1) a useful tool in the diagnosis of AH/EIN in EMP; (2) PAX2 loss should be interpreted with caution and in combination with morphology and other markers.

Correlation of cone biopsy with findings at radical hysterectomy and use of adjuvant radiation therapy.

OBJECTIVE: To determine if pathologic findings in cone biopsy specimens correlate with residual invasive disease in radical hysterectomy specimens and the need for adjuvant chemo-radiation therapy. STUDY DESIGN: We identified 65 patients who underwent a cone biopsy and subsequent radical hysterectomy. Clinico-pathologic parameters in the cone specimens were correlated with the presence of residual invasive disease in the radical hysterectomy specimens and the need for adjuvant chemo-radiation. RESULTS: A positive endocervical margin, a positive deep margin, a positive post-cone ECC, and positive LVSI were significantly associated with the presence of residual disease in the radical hysterectomy specimen, while positive LVSI, a positive ECC, a positive deep cone margin, and greater than 1 positive margin were significantly associated with the use of adjuvant chemo-radiation therapy. CONCLUSION: Pathologic parameters in cone biopsy specimens can estimate the risk of residual invasive disease in radical hysterectomy specimens and the use of adjuvant chemo-radiation.

Morules But Not Squamous Differentiation are a Reliable Indicator of CTNNB1 (ß-catenin) Mutations in Endometrial Carcinoma and Precancers.

Although collectively regarded as "squamous differentiation (SD)" in endometrial endometrioid carcinoma (EEC) and atypical hyperplasia/endometrioid intraepithelial neoplasia (AH/EIN), morules (often referred to as "squamous morules") and true SD may represent two distinct phenomena. Here, we explored the distinction between morules versus SD and investigated the association of morules and SD with CTNNB1 mutations. A total of 270 cases of EEC and AH/EIN were studied, including EEC with (n=36) or without (n=36) morules and AH/EIN with (n=80) or without (n=118) morules. Cases were analyzed by immunohistochemistry and selected cases (n=20) by targeted next-generation sequencing panel. Near-perfect agreement was found between morules and glandular ß-catenin nuclear staining in AH/EIN and EEC. A strong positive association was found between morules and glandular ß-catenin nuclear staining ( P <0.0001, Φ=0.59 in AH/EIN; P <0.0001, Φ=0.85 in EEC). There was no association between (1) morules and glandular PAX2 or PTEN aberrant expression or (2) SD and aberrant expression of ß-catenin, PAX2 or PTEN (Φ=0.09, ß-catenin; Φ=0.16, PAX2; Φ=0.13, PTEN). CTNNB1 mutations were identified in all 20 selected morule-containing cases (100%). Next-generation sequencing was performed on 2 (preprogestin and postprogestin treatment) biopsies from 1 patient, revealing identical mutational profile in morules and glands. In conclusion, (1) SD and morules are distinct biological phenomena; (2) the presence of morules, but not SD, is a reliable indicator of CTNNB1 mutations in EEC and AH/EIN. Our findings demonstrate that SD and morules are distinct biological phenomena. Since morules but not SD are associated with ß-catenin mutations, the distinction is clinically relevant and should be included in diagnostic reports.

Reliable Identification of Endometrial Precancers Through Combined Pax2, ß-Catenin, and Pten Immunohistochemistry.

The diagnosis of endometrial atypical hyperplasia/endometrioid intraepithelial neoplasia (AH/EIN) remains challenging and subjective in some cases, with variable histologic criteria and differences of opinion among gynecologic pathologists, potentially leading to under/overtreatment. There has been growing interest in the use of specific immunohistochemical markers as adjuncts in AH/EIN diagnosis. For example, the World Health Organization 2020 Classification specifies that loss of Pten, Pax2, or mismatch repair proteins are desirable diagnostic criteria. Other markers, most notably ß-catenin and Arid1a, are also aberrantly expressed in some AH/EIN. However, the performance of some markers individually-and more importantly as a group-has not been rigorously explored, raising questions as to which marker(s) or combination(s) is the most effective in practice. Formalin-fixed paraffin-embedded tissue sections from AH/EIN cases (n=111) were analyzed by immunohistochemistry for 6 markers: Pax2, Pten, Mlh1, ß-catenin, Arid1a, and p53. Aberrant expression was tabulated for each case and marker. An additional set of normal endometria (n=79) was also analyzed to define optimal diagnostic criteria for marker aberrance. The performance characteristics of each marker, the entire panel, and subsets thereof were quantitatively and statistically analyzed. In order of number of cases detected, the most frequently aberrant markers in AH/EIN were Pax2 (81.1% of cases), Pten (50.5%), ß-catenin (47.7%), Arid1a (7.2%), Mlh1 (4.5%), and p53 (2.7%). The majority of cases showed aberrant expression of ≥2 markers. All 6 markers together identified 92.8% of cases. Arid1a, Mlh1, and p53 were robust and readily scored markers, but all cases showing aberrant expression of these 3 markers were also detected by Pax2, Pten, or ß-catenin. A focused panel of only 3 markers (Pax2, Pten, and ß-catenin) showed optimal performance characteristics as a diagnostic adjunct in the histopathologic diagnosis of AH/EIN. Use of this panel is practicable and robust, with at least 1 of the 3 markers being aberrant in 92.8% of AH/EIN.

Significance of Oil-Red-O positive macrophages in bronchoalveolar lavage in diagnosing E-cigarettes or vaping product use-associated lung injury: A case series.

BACKGROUND: Lipid-laden macrophages detected by Oil-Red-O (ORO) stain in fresh bronchoalveolar lavage (BAL) specimens have been proposed as a potential diagnostic marker for E-cigarettes or vaping product use-associated lung injury (EVALI). However, studies are few, and the sensitivity and specificity of the test have not been thoroughly investigated. METHODS: We performed ORO stain on fresh BAL specimens from six confirmed EVALI and 36 non-EVALI patients. After semi-quantitative analysis, the sensitivity and specificity of ORO-positive macrophages (OPM) for detection of EVALI were calculated. RESULTS: No significant difference in cytomorphology or raw macrophage count was observed between EVALI and non-EVALI groups (49% vs 55% of all nucleated cells). However, with ORO stain, all EVALI specimens (6/6) showed a high percentage (≥50% of all macrophages) of OPM (mean 87%), and large (≥25% of host macrophage nuclear size) lipid droplets (mean 42%), while the majority of non-EVALI specimens showed a low percentage of OPM (32/36, mean 10%), and small lipid droplets (34/36, mean 6%). The differences between the two groups in both high OPM and large lipid droplet rates are statistically significant (P < .0001 for both comparisons). The combined sensitivity and specificity of high OPM and large lipid droplets for diagnosing EVALI were 100% and 94%, respectively. CONCLUSION: In BAL specimens obtained from patients with clinically suspected EVALI, a high percentage of OPM with large lipid droplets showed high sensitivity and specificity for the diagnosis of EVALI and may serve as a potentially useful tool in the evaluation of vaping-related lung injury, improving diagnostic accuracy.

PD-L1 Expression in Endocervical Adenocarcinoma: Correlation With Patterns of Tumor Invasion, CD8+ Tumor-infiltrating Lymphocytes, and Clinical Outcomes.

Programmed death-1 ligand (PD-L1) expression has been used as a predictive marker for response to immune checkpoint inhibitors and has been reported to have prognostic value. Its prevalence and significance in endocervical adenocarcinoma (ECA) remain underinvestigated. We evaluated PD-L1 expression and CD8+ tumor-infiltrating lymphocyte density in whole tissue sections of 89 ECAs. PD-L1 expression was observed in 68% of ECAs by combined positive score (CPS, cutoff 1) and 29% of ECAs by tumor proportion score (TPS, cutoff 1%). Using CPS, PD-L1 expression was seen in 11%, 78%, and 72% of pattern A, B, and C tumors, respectively, with significantly higher expression in tumors with destructive-type invasion (B and C) (P=0.001 [A vs. B], 0.0006 [A vs. C], 0.0002 [A vs. B+C]). Using TPS, no significant difference in PD-L1 expression was seen between tumors with different invasion patterns (0%, 22%, and 32% in tumors with pattern A, B, and C, respectively; P=0.27 [A vs. B], 0.053 [A vs. C], 0.11 [A vs. B+C]). PD-L1-positive ECAs demonstrated significantly higher CD8+ tumor-infiltrating lymphocyte density (CPS: P=0.028; TPS: P=0.013) and worse progression-free survival when compared with PD-L1-negative ECAs (CPS: hazard ratio [HR]=4.253 vs. 0.235, P=0.025; TPS: HR=4.98 vs. 0.2; P=0.004). When invasion patterns were separately assessed, pattern C tumors similarly showed worse progression-free survival in PD-L1-positive tumors (CPS: HR=6.15 vs. 0.16, P=0.045; TPS: HR=3.78 vs. 0.26, P=0.027). In conclusion, our data show frequent PD-L1 expression in ECA with destructive-type invasion, supporting the role of the PD-1/PD-L1 pathway as a therapeutic target for these tumors. Our data also support PD-L1 as a negative prognostic marker associated with a potentially unfavorable outcome.

A Surgical Window Trial Evaluating Medroxyprogesterone Acetate with or without Entinostat in Patients with Endometrial Cancer and Validation of Biomarkers of Cellular Response.

PURPOSE: This surgical window of opportunity (window) study assessed the short-term effect of medroxyprogesterone acetate (MPA) alone versus MPA plus the histone deacetylase (HDAC) inhibitor entinostat on regulation of progesterone receptor (PR) in women with newly diagnosed endometrioid endometrial adenocarcinoma. PATIENTS AND METHODS: This multisite, randomized, open-label surgical window study treated women intramuscularly on day 1 with 400 mg MPA. Entinostat given 5 mg by mouth on days 1, 8, and 15 was randomly assigned with equal probability. Surgery followed on days 21-24. Pretreatment and posttreatment tissue was assessed for PR H-scores, Ki-67 levels, and histologic response. RESULTS: Fifty patients were accrued in 4 months; 22 and 20 participants had PR evaluable pretreatment and posttreatment slides in the MPA and MPA/entinostat arms, respectively. Median posttreatment PR H-scores were significantly lower than pretreatment H-scores in both arms but did not differ significantly (MPA: 247 vs. 27, MPA/entinostat 260 vs. 23, respectively, P = 0.87). Decreased Ki-67 was shown in 90% treated with MPA/entinostat compared with 68% treated with MPA alone (P = 0.13). Median PR H-score decreases were larger when Ki-67 was decreased (208) versus not decreased (45). The decrease in PR pretreatment versus posttreatment was associated with loss of Ki-67 nuclear staining, consistent with reduced cellular proliferation (P < 0.008). CONCLUSIONS: This surgical window trial rapidly accrued in a multisite setting and evaluated PR as its primary endpoint and Ki-67 as secondary endpoint. Despite no immediate effect of entinostat on PR in this short-term study, lessons learned can inform future window and treatment trials.