Lesions sub-diagnostic of endometrioid intra-epithelial neoplasia/atypical hyperplasia: value of morphology and immunohistochemistry in predicting neoplastic outcome.

AIMS: Areas of gland crowding that do not fulfil diagnostic criteria of endometrioid intra-epithelial neoplasia (EIN) are often encountered in endometrial biopsies. In this study, we document the prevalence of neoplastic outcome in patients with these subdiagnostic lesions (SL) and assess the utility of morphological features and a three-marker immunohistochemistry panel (PAX2, PTEN, beta-catenin) to predict outcome. METHODS AND RESULTS: Of 430 women with SL on endometrial sampling at Brigham and Women's Hospital between 2001 and 2021 with available follow-up biopsy, 72 (17%) had a neoplastic outcome (EIN or endometrioid carcinoma). Multilayered epithelium and mitoses in SL were statistically associated with a neoplastic outcome. Abnormal three-marker staining was observed in 93% (53 of 57) of SL with neoplastic outcome and 60% (37 of 62) of a control group with benign outcome. Among the 72 patients with neoplastic outcome, EIN/carcinoma tissue was available in 33; of these, 30 (91%) showed abnormal staining for one or more markers. Remarkably, in 84% of these cases the EIN/carcinoma had the aberrant expression seen in the preceding SL. Based on a prevalence of 17%, the positive and negative predictive values of abnormal staining in one or more markers were 24 and 97%, respectively. CONCLUSIONS: The presence of SL warrants clinical surveillance and repeat sampling because it is followed by endometrioid neoplasia in a significant subset of patients. Normal three-marker staining identifies women with a very low risk of neoplastic outcome. Conversely, abnormal staining is frequent in SL with benign outcome leading to poor specificity and positive predictive value.

Localized Endometrial Proliferations of Pregnancy are Clonal Glandular Outgrowths Characterized by PTEN Loss and PIK3CA Pathogenic Variants.

Gestational endometrium can demonstrate a spectrum of atypical but benign changes. One such lesion is localized endometrial proliferation of pregnancy (LEPP), first described in a series of 11 cases. To understand its biological and clinical importance, we explore the pathologic, immunophenotypic, and molecular features of this entity. Nine cases of LEPP identified in 15 years were retrieved from departmental archives and reviewed. Immunohistochemistry and next-generation sequencing using a comprehensive 446-gene panel were performed when the material was available. Eight cases were identified in curettage specimens performed after first-trimester pregnancy loss, and 1 in the basal plate of a mature placenta. The mean patient age was 35 (range 27-41) years. The mean lesion size was 6.3 (range 2-12) mm. Architectural patterns, often coexisting in the same case, included cribriform (n = 7), solid (n = 5), villoglandular (n = 2), papillary (n = 2), and micropapillary (n = 1). Cytologic atypia was mild in 7 cases and moderate in 2. Mitotic activity was low (up to 3 per 2.4 mm2). All lesions were associated with neutrophils. Background Arias-Stella phenomenon was present in 4 cases. Immunohistochemistry was performed in 7 LEPP, all of which demonstrated wildtype p53, retained MSH6 and PMS2, membranous beta-catenin, and positive estrogen receptor (mean 71%) and progesterone receptor (mean 74%). All were negative for p40 except 1 case (focal weak positivity). PTEN was markedly reduced in background secretory glands in all cases; in 5/7, LEPP foci showed a complete absence of PTEN expression. PIK3CA pathogenic variants were identified in 4/4 cases sequenced; 3/4 had inactivating PTEN mutations. Follow-up, available in 8 patients (mean length = 51 months, range 7-161), was conservative with observation only and showed no persistence or adverse outcomes. LEPP is characterized by intraglandular cribriform/solid architecture, positive estrogen receptor/progesterone receptor, PTEN loss, and PIK3CA and PTEN mutations. Although our findings indicate that LEPP is neoplastic, for now, we advise against diagnosing LEPP as endometrial carcinoma or hyperplasia because LEPP has a particular clinicopathologic context (concurrent gestation), distinct morphology (purely intraepithelial complex growth), and indolent outcome. Thus, it should be distinguished from endometrial intraepithelial neoplasia and carcinoma for which therapeutic interventions are indicated.

Computational augmentation of neoplastic endometrial glands in digital pathology displays.

The pathologic diagnosis of neoplasia requires localization and classification of lesional tissue, a process that depends on the recognition of an abnormal spatial distribution of neoplastic elements relative to admixed normal background tissue. In endometrial intraepithelial neoplasia (EIN), a pre-cancer usually managed by hysterectomy, a clonally mutated proliferation of cytologically altered glands ('neoplastic-EIN') aggregates in clusters that also contain background non-neoplastic glands ('background-NL'). Here, we used image analysis to classify individual glands within endometrial tissue fragments as neoplastic-EIN or background-NL, and we used the distribution of predictions to localize foci diagnostic of EIN. Nuclear coordinates were automatically assigned and were used as vertices to generate Delaunay triangulations for each gland. Graph statistical variables were used to develop random forest algorithms to classify glands as neoplastic-EIN or background-NL. Individual glands in an independent validation set were scored by a 'ground truth' biomarker (PAX2 immunohistochemistry). We found that exclusion of small glands led to improvement in classification accuracy. Using an inclusion threshold of 200 nuclei per gland, our final model classification accuracy was 77.5% in the validation set, with a positive predictive value of 0.81. We leveraged this high positive predictive value in a point cloud overlay display to assist end-user identification of EIN foci. This study demonstrates that graph theory approaches applied to small-scale anatomic elements in the endometrium allow biologic classification by machine learning, and that spatial superimposition over large-scale tissue expanses can have practical diagnostic utility. We expect this augmented diagnostic approach to be generalizable to commonly encountered problems in other organ systems. © 2020 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Deciphering serous ovarian carcinoma histopathology and platinum response by convolutional neural networks.

BACKGROUND: Ovarian cancer causes 151,900 deaths per year worldwide. Treatment and prognosis are primarily determined by the histopathologic interpretation in combination with molecular diagnosis. However, the relationship between histopathology patterns and molecular alterations is not fully understood, and it is difficult to predict patients' chemotherapy response using the known clinical and histological variables. METHODS: We analyzed the whole-slide histopathology images, RNA-Seq, and proteomics data from 587 primary serous ovarian adenocarcinoma patients and developed a systematic algorithm to integrate histopathology and functional omics findings and to predict patients' response to platinum-based chemotherapy. RESULTS: Our convolutional neural networks identified the cancerous regions with areas under the receiver operating characteristic curve (AUCs) > 0.95 and classified tumor grade with AUCs > 0.80. Functional omics analysis revealed that expression levels of proteins participated in innate immune responses and catabolic pathways are associated with tumor grade. Quantitative histopathology analysis successfully stratified patients with different response to platinum-based chemotherapy (P = 0.003). CONCLUSIONS: These results indicated the potential clinical utility of quantitative histopathology evaluation in tumor cell detection and chemotherapy response prediction. The developed algorithm is easily extensible to other tumor types and treatment modalities.

A New Classification of Benign, Premalignant, and Malignant Endometrial Tissues Using Machine Learning Applied to 1413 Candidate Variables.

Benign normal (NL), premalignant (endometrial intraepithelial neoplasia, EIN) and malignant (cancer, EMCA) endometria must be precisely distinguished for optimal management. EIN was objectively defined previously as a regression model incorporating manually traced histologic variables to predict clonal growth and cancer outcomes. Results from this early computational study were used to revise subjective endometrial precancer diagnostic criteria currently in use. We here use automated feature segmentation and updated machine learning algorithms to develop a new classification algorithm. Endometrial tissue from 148 patients was randomly separated into 72-patient training and 76-patient validation cohorts encompassing all 3 diagnostic classes. We applied image analysis software to keratin stained endometrial tissues to automatically segment whole-slide digital images into epithelium, cells, and nuclei and extract corresponding variables. A total of 1413 variables were culled to 75 based on random forest classification performance in a 3-group (NL, EIN, EMCA) model. This algorithm correctly classifies cases with 3-class error rates of 0.04 (training set) and 0.058 (validation set); and 2-class (NL vs. EIN+EMCA) error rate of 0.016 (training set) and 0 (validation set). The 4 most heavily weighted variables are surrogates of those previously identified in manual-segmentation machine learning studies (stromal and epithelial area percentages, and normalized epithelial surface lengths). Lesser weighted predictors include gland and lumen axis lengths and ratios, and individual cell measures. Automated image analysis and random forest classification algorithms can classify normal, premalignant, and malignant endometrial tissues. Highest predictive variables overlap with those discovered independently in early models based on manual segmentation.

Issues in the Differential Diagnosis of Uterine Low-grade Endometrioid Carcinoma, Including Mixed Endometrial Carcinomas: Recommendations from the International Society of Gynecological Pathologists.

This article provides practical recommendations developed from the International Society of Gynecological Pathologists Endometrial Carcinoma Project to address 4 issues that may arise in the diagnosis of uterine corpus low-grade endometrioid carcinoma: (1) The distinction between atypical hyperplasia and low-grade endometrioid carcinoma. (2) The distinction between low-grade endometrioid carcinoma and serous carcinoma. (3) The distinction between corded and hyalinized or spindle cell variants of low-grade endometrioid carcinoma and carcinosarcoma. (4) The diagnostic criteria for mixed endometrial carcinomas, a rare entity that should be diagnosed only after exclusion of a spectrum of tumors including morphologic variants of endometrioid carcinoma, dedifferentiated endometrial carcinoma, carcinosarcoma, and endometrial carcinomas with ambiguous morphology.

Assessment of a Chemotherapy Response Score (CRS) System for Tubo-Ovarian High-Grade Serous Carcinoma (HGSC).

A chemotherapy response score (CRS) system was recently described to assess the histopathologic response and prognosis of patients with tubo-ovarian high-grade serous carcinoma (HGSC) receiving neoadjuvant chemotherapy. The current study was performed as an independent assessment of this CRS system. We retrospectively identified advanced stage HGSC patients who received neoadjuvant chemotherapy and underwent interval debulking. If available, a hemotoxylin and eosin slide from the omentum and the adnexa was selected for the study. Slides were independently scored by 13 pathologists using the 3-tiered CRS system. Reviewers then received web-based training and rescored the slides. Overall survival and progression-free survival were estimated using the Kaplan-Meier method and compared using the log-rank test. A total of 68 patients with omental (n=65) and/or adnexal (n=59) slides were included in the study. Interobserver reproducibility was moderate for omentum (κ, 0.48) and poor for adnexa (κ, 0.40), which improved for omentum (κ, 0.62) but not for adnexa (κ, 0.38) after online training. For omental slides, a consensus CRS of 1/2 was associated with a shorter median progression-free survival (10.9 mo; 95% confidence interval, 9-14) than a CRS of 3 (18.9 mo; 95% CI, 18-24; P=0.020). In summary, a 3-tiered CRS system of hemotoxylin and eosin-stained omental deposits can yield prognostic information for HGSC patients receiving neoadjuvant chemotherapy, and web-based training improved reproducibility but did not alter determination of clinical outcomes. The CRS system may allow oncologists to identify potential nonresponders and triage HGSC patients for heightened observation and/or clinical trials.

Biomarker resolution of uterine smooth muscle tumor necrosis as benign vs malignant.

Uterine leiomyosarcomas are rare malignant tumors with a poor prognosis while leiomyomas are common benign tumors unrelated to their malignant counterparts. Diagnostic features commonly present in leiomyosarcoma include cytologic atypia, high mitotic index, and a sarcoma-specific geographic cell death designated 'tumor cell necrosis (TCN)'. TCN has a sharp viable-nonviable boundary lacking inflammation, fibrosis or granulation tissue seen in nonspecific infarction. These characteristics are sometimes difficult to interpret on routine hematoxylin and eosin slides, and can lead to diagnostic errors. In this study, we used extracellular matrix stains to test the hypothesis that the host response which characterizes nonspecific infarction may degrade the matrix in infarcted tumor more than in TCN. A 'honeycomb' pattern of reticulin highlighted individual tumor cells in viable regions of all cases. Nonviable area of reticulin patterns differed significantly by diagnosis (P<0.001), with a honeycomb pattern maintained (91%, 20/22) in leiomyosarcoma and lost (61%, 11/18) in leiomyomas. Retention of honeycomb reticulin in nonviable areas of leiomyosarcoma occurred irrespective of the presence of inflammation, hemorrhage, fibrosis, or diffuse hyalinization. Fibrosis/hyalinization as evidenced by trichrome stain was significantly (P<0.001) more common in nonviable areas of benign leiomyomas (100%, 18/18) compared with leiomyosarcomas (36%, 8/22). In those occasions where viable tissues contained discernable polarization of mitotic activity, these decreased toward the nonviable interface in leiomyosarcoma, and had an opposite pattern in leiomyomas, increasing toward the interface. There is a significant difference in the reticulin and collagen networks of nonviable areas of leiomyosarcoma compared with leiomyoma. At the time of early injury, both retain reticulin; however, this is cleared over time in benign, but not malignant, areas of necrosis. We conclude that proliferative repair of leiomyomas at the viable-nonviable interface includes remodeling of the extracellular matrix, in contrast to the static preservation of extracellular matrix ('mummification') in nonviable areas of leiomyosarcomas.

Dyssynchronous secretory endometrial glands often show sporadically acquired progesterone nonresponsiveness.

Primary sporadic gene-inactivating events within the progesterone response cascade might explain the presence of individual dyssynchronous (outlier) glands commonly observed in a secretory background. We queried morphologically dyssynchronous glands in mid-secretory endometrium with a series of markers normally downregulated by progesterone. Seventy-nine mid-secretory endometrial biopsies were stained with hematoxylin and eosin, MIB-1, PAX2, estrogen and progesterone receptors, and PTEN. Aberrant staining of glands was independently scored for each marker. Outlier glands overlapping between stains were enumerated. A total of 63% of cases had hematoxylin and eosin stained outlier glands (average 9), which often demonstrated failed progesterone-mediated downregulation of PAX2 (43%), estrogen (40%), and/or progesterone receptors (28%). Aberrations of progesterone response was seen in 70% to 85% of cases overall, averaging 10 to 30 glands/affected case. The frequency and burden of affected glands was similar to that seen for primary inactivating events of the PAX2 and PTEN genes (35% and 41% of cases, respectively, averaging 32 and 38 glands per affected patient). Sporadic gene-inactivating events are common during endometrial regeneration, and may cause morphologic changes unmasked by the hormonal context. Some of these dyssynchronous "outlier" glands, whether evident on hematoxylin and eosin stain or not, have an interrupted progesterone response.

Hypertension and Risk of Endometrial Cancer: A Pooled Analysis in the Epidemiology of Endometrial Cancer Consortium (E2C2).

BACKGROUND: The incidence rates of endometrial cancer are increasing, which may partly be explained by the rising prevalence of obesity, an established risk factor for endometrial cancer. Hypertension, another component of metabolic syndrome, is also increasing in prevalence, and emerging evidence suggests that it may be associated with the development of certain cancers. The role of hypertension independent of other components of metabolic syndrome in the etiology of endometrial cancer remains unclear. In this study, we evaluated hypertension as an independent risk factor for endometrial cancer and whether this association is modified by other established risk factors. METHODS: We included 15,631 endometrial cancer cases and 42,239 controls matched on age, race, and study-specific factors from 29 studies in the Epidemiology of Endometrial Cancer Consortium. We used multivariable unconditional logistic regression models to estimate ORs and 95% confidence intervals (CI) to evaluate the association between hypertension and endometrial cancer and whether this association differed by study design, race/ethnicity, body mass index, diabetes status, smoking status, or reproductive factors. RESULTS: Hypertension was associated with an increased risk of endometrial cancer (OR, 1.14; 95% CI, 1.09-1.19). There was significant heterogeneity by study design (Phet < 0.01), with a stronger magnitude of association observed among case-control versus cohort studies. Stronger associations were also noted for pre-/perimenopausal women and never users of postmenopausal hormone therapy. CONCLUSIONS: Hypertension is associated with endometrial cancer risk independently from known risk factors. Future research should focus on biologic mechanisms underlying this association. IMPACT: This study provides evidence that hypertension may be an independent risk factor for endometrial cancer.

Endometrial intraepithelial neoplasia with secretory differentiation: diagnostic features and underlying mechanisms.

Endometrial intraepithelial neoplasia (EIN) with secretory differentiation and ordinary EIN occurring in a secretory context are rare but recognized findings. We determined how often secretory differentiation in EIN was associated with evidence of circulating progestins in the background endometrium, and studied clinical characteristics and clinical outcomes of affected patients. We selected 41 patients with secretory differentiation in either the EIN itself (n=31) and/or background endometrium (n=38). Most (90%, 28/31) secretory EINs were associated with circulating progestins. Rare exceptions were observed, suggesting that secretory EIN may occur as a hormone-independent phenomenon. Circulating progestins are not sufficient, however, to induce EIN secretory differentiation, as 26% (10/38) of EIN within a secretory background were of the ordinary (non-secretory) type. EIN patients with secretory endometrium in the background are younger (averaging 45 years) than the aggregate group of all patients with EIN (53 years in previously published studies) and are often premenopausal with a cyclical source of endogenous progestins. Involution of EIN during follow-up was more frequent (81%, 17/21) for those with a secretory background at the time of initial EIN diagnosis compared with historical averages (25%, 36/142). These results suggest a potential role for endogenous progesterone, as well as therapeutic progestins, in modulating EIN outcomes.

Reproducibility of endometrial intraepithelial neoplasia diagnosis is good, but influenced by the diagnostic style of pathologists.

Endometrial intraepithelial neoplasia (EIN) applies specific diagnostic criteria to designate a monoclonal endometrial preinvasive glandular proliferation known from previous studies to confer a 45-fold increased risk for endometrial cancer. In this international study we estimate accuracy and precision of EIN diagnosis among 20 reviewing pathologists in different practice environments, and with differing levels of experience and training. Sixty-two endometrial biopsies diagnosed as benign, EIN, or adenocarcinoma by consensus of two expert subspecialty pathologists were used as a reference comparison to assess diagnostic accuracy of 20 reviewing pathologists. Interobserver reproducibility among the 20 reviewers provided a measure of diagnostic precision. Before evaluating cases, observers were self-trained by reviewing published textbook and/or online EIN diagnostic guidelines. Demographics of the reviewing pathologists, and their impressions regarding implementation of EIN terminology were recorded. Seventy-nine percent of the 20 reviewing pathologists' diagnoses were exactly concordant with the expert consensus (accuracy). The interobserver weighted κ values of 3-class EIN scheme (benign, EIN, carcinoma) diagnoses between expert consensus and each of reviewing pathologists averaged 0.72 (reproducibility, or precision). Reviewing pathologists demonstrated one of three diagnostic styles, which varied in the repertoire of diagnoses commonly used, and their nonrandom response to potentially confounding diagnostic features such as endometrial polyp, altered differentiation, background hormonal effects, and technically poor preparations. EIN diagnostic strategies can be learned and implemented from standard teaching materials with a high degree of reproducibility, but is impacted by the personal diagnostic style of each pathologist in responding to potential diagnostic confounders.

Measuring Digital Pathology Throughput and Tissue Dropouts.

BACKGROUND: Digital pathology operations that precede viewing by a pathologist have a substantial impact on costs and fidelity of the digital image. Scan time and file size determine throughput and storage costs, whereas tissue omission during digital capture ("dropouts") compromises downstream interpretation. We compared how these variables differ across scanners. METHODS: A 212 slide set randomly selected from a gynecologic-gestational pathology practice was used to benchmark scan time, file size, and image completeness. Workflows included the Hamamatsu S210 scanner (operated under default and optimized profiles) and the Leica GT450. Digital tissue dropouts were detected by the aligned overlay of macroscopic glass slide camera images (reference) with images created by the slide scanners whole slide images. RESULTS: File size and scan time were highly correlated within each platform. Differences in GT450, default S210, and optimized S210 performance were seen in average file size (1.4 vs. 2.5 vs. 3.4 GB) and scan time (93 vs. 376 vs. 721 s). Dropouts were seen in 29.5% (186/631) of successful scans overall: from a low of 13.7% (29/212) for the optimized S210 profile, followed by 34.6% (73/211) for the GT450 and 40.4% (84/208) for the default profile S210 profile. Small dislodged fragments, "shards," were dropped in 22.2% (140/631) of slides, followed by tissue marginalized at the glass slide edges, 6.2% (39/631). "Unique dropouts," those for which no equivalent appeared elsewhere in the scan, occurred in only three slides. Of these, 67% (2/3) were "floaters" or contaminants from other cases. CONCLUSIONS: Scanning speed and resultant file size vary greatly by scanner type, scanner operation settings, and clinical specimen mix (tissue type, tissue area). Digital image fidelity as measured by tissue dropout frequency and dropout type also varies according to the tissue type and scanner. Dropped tissues very rarely (1/631) represent actual specimen tissues that are not represented elsewhere in the scan, so in most cases cannot alter the diagnosis. Digital pathology platforms vary in their output efficiency and image fidelity to the glass original and should be matched to the intended application.

Secretory cell outgrowth, PAX2 and serous carcinogenesis in the Fallopian tube.

The 'p53 signature' is a benign secretory cell outgrowth in the distal Fallopian tube that shares properties with ovarian serous cancer-including p53 mutations-and is a putative serous cancer precursor. We expanded the precursor definition to all secretory cell outgrowths (SCOUTs) of 30 or more cells and scored normal (N) and altered (A) expression of both p53 and PAX2, a gene down-regulated in ovarian and endometrial cancer. SCOUTs were identified by BCL2/p73 staining in tubes from women with serous carcinoma, inherited mutations in BRCA1 or BRCA2 and controls. SCOUTs were prevalent in both proximal and distal tube and significantly associated with serous carcinoma versus the others (p < 0.001); 89% were PAX2 (A) and 26% were PAX2 (A)/p53 (A) (p53 signatures). PAX2 (A)/p53 (N) SCOUTs were free of p53 mutations; however, 12 of 13 p53 signatures were PAX2 (A). A tubal carcinoma and contiguous SCOUT were p53 (A)/PAX2 (A) and shared the same p53 mutation. SCOUTs are discretely localized alterations commonly containing altered expression of multiple genes within histologically benign tubal epithelium. Geographic distribution in the tube varies by genotype and immunophenotype, from regionally unrestricted (PAX2) to greater likelihood specific area (fimbria) of shared prevalence (PAX2 and p53). This study reveals, for the first time, an entity (SCOUT) that is associated with serous cancer, expands the topography of altered PAX2 expression in the female genital tract mucosa and highlights another potential pathway disturbance involved in early serous carcinogenesis in the Fallopian tube.

Endometrial Tumor Classification by Histomorphology and Biomarkers in the Nurses' Health Study.

OBJECTIVE: Endometrial cancers have historically been classified by histomorphologic appearance, which is subject to interobserver disagreement. As molecular and biomarker testing has become increasingly available, the prognostic significance and accuracy of histomorphologic diagnoses have been questioned. To address these issues for a large, prospective cohort study, we provide the results of a centralized pathology review and biomarker analysis of all incidental endometrial carcinomas occurring between 1976 and 2012 in the Nurses' Health Study. METHODS: Routine histology of all (n = 360) cases was reviewed for histomorphologic diagnosis. Cases were subsequently planted in a tissue microarray to explore expression of a variety of biomarkers (e.g., ER, PR, p53, PTEN, PAX2, AMACR, HNF1ß, Napsin A, p16, PAX8, and GATA3). RESULTS: Histologic subtypes included endometrioid (87.2%), serous (5.6%), carcinosarcoma (3.9%), clear cell (1.7%), and mixed type (1.7%). Biomarker results within histologic subtypes were consistent with existing literature: abnormal p53 was frequent in serous cases (74%), and HNF1ß (67%), Napsin A (67%), and AMACR (83%) expression was frequent in clear cell carcinomas. Our dataset also allowed for examination of biomarker expression across non-preselected histologies. The results demonstrated that (1) HNF1ß was not specific for clear cell carcinoma, (2) TP53 mutations occurred across many histologies, and (3) GATA3 was expressed across multiple histotypes, with 75% of positive cases demonstrating high-grade features. CONCLUSIONS: Our findings establish the subtypes of endometrial cancer occurring in the Nurses' Health Study, corroborate the sensitivity of certain well-established biomarkers, and call into question previously identified associations between certain biomarkers (e.g., HNF1B) and particular histotypes.

Breast-cancer stromal cells with TP53 mutations and nodal metastases.

BACKGROUND: The importance of cross-talk between a cancer and its microenvironment has been increasingly recognized. We hypothesized that mutational inactivation of the tumor-suppressor gene TP53 and genomic alterations in stromal cells of a tumor's microenvironment contribute to the clinical outcome. METHODS: We performed TP53 mutation analysis and genomewide analysis of loss of heterozygosity and allelic imbalance on DNA from isolated neoplastic epithelial and stromal cells from 43 patients with hereditary breast cancer and 175 patients with sporadic breast cancer. Compartment-specific patterns and TP53 mutations were analyzed. Associations between compartment-specific TP53 status, loss of heterozygosity or allelic imbalance, and clinical and pathological characteristics were computed. RESULTS: TP53 mutations were associated with an increased loss of heterozygosity and allelic imbalance in both hereditary and sporadic breast cancers, but samples from patients with hereditary disease had more frequent mutations than did those from patients with sporadic tumors (74.4% vs. 42.3%, P=0.001). Only 1 microsatellite locus (2p25.1) in stromal cells from hereditary breast cancers was associated with mutated TP53, whereas there were 66 such loci in cells from sporadic breast cancers. Somatic TP53 mutations in stroma, but not epithelium, of sporadic breast cancers were associated with regional nodal metastases (P=0.003). A specific set of five loci linked to an increased loss of heterozygosity and allelic imbalance in the stroma of sporadic tumors was associated with nodal metastases in the absence of TP53 mutations. No associations were found between any of the clinical or pathological features of hereditary breast cancer with somatic TP53 mutations. CONCLUSIONS: Stroma-specific loss of heterozygosity or allelic imbalance is associated with somatic TP53 mutations and regional lymph-node metastases in sporadic breast cancer but not in hereditary breast cancer.

Clinical outcome in diagnostically ambiguous foci of 'gland crowding' in the endometrium.

Premalignant endometrial lesions (endometrial intraepithelial neoplasia (EIN)) are clonal neoplasms that arise focally and can be diagnosed using specific criteria: (1) area of glands exceeds that of stroma (glands/stroma >1), (2) nuclear and/or cytoplasmic features of epithelial cells differ between architecturally abnormal glands and normal background glands, and (3) maximum linear dimension exceeds 1 mm. However, localized groups of crowded endometrial glands may be encountered that do not fulfill all of the criteria for EIN, are interpreted as ambiguous, and are reported as 'focal gland crowding'. We conducted a retrospective study of gland crowding using a free-text index search for this term in our pathology files. The age of the patients, number of subsequent specimens, the duration, and the outcome of the follow-ups were recorded. Of the 71,579 consecutive gynecological pathology reports, 206 (0.3%) 'gland crowding' cases were identified, in which 69% (143/206) had follow-up sampling. Of these, 33 (23%) had an outcome diagnosis of EIN (27 cases; 19%) or carcinoma (6 cases; 4%). Included were 18 cases (55%) diagnosed within the first year and presumed concurrent, and an additional 15 (45%) discovered after 1 year and interpreted as a later phase of disease or new events. The term 'crowded glands' is a highly significant finding that carries a substantial risk of an outcome of EIN and occasionally malignancy. It underscores the importance of follow-up when some but not all of the criteria for EIN are encountered in the appropriate clinical setting.

High-grade fimbrial-ovarian carcinomas are unified by altered p53, PTEN and PAX2 expression.

High-grade endometrioid and serous carcinomas of the ovary and fallopian tube are responsible for the majority of cancer deaths and comprise a spectrum that includes early or localized (tubal intraepithelial carcinoma) and advanced (invasive or metastatic) disease. We subdivided a series of these tumors into three groups, (1) classic serous, (2) mixed serous and endometrioid and (3) endometrioid carcinomas and determined: (1) the frequencies of coexisting tubal intraepithelial carcinoma, (2) frequency of a dominant ovarian mass suggesting an ovarian origin and (3) immuno-localization of WT-1, p53, PTEN, PAX2 and p16(ink4). All tumors were analyzed for p53 mutations. Thirty six, 25 and 8% of groups 1-3 were associated with tubal intraepithelial carcinoma (P=0.09) and 34, 45 and 62% predominated in one ovary (P=0.028), respectively. Differences in frequencies of diffuse p53 immunostaining (85-93%), WT-1 (70-98%) and p16(ink4) positivity (69-75%) were not significant for all groups. Greater than 95% reduction in PAX2 and PTEN occurred in 67-75 and 5-12%, respectively; however, PAX2 and PTEN staining intensity, when present, was often heterogeneous, highlighting different tumor populations. PAX2 and PTEN expression were markedly reduced or absent in 12 of 12 and 4 of 12 tubal intraepithelial carcinomas. In summary, high-grade müllerian carcinomas share identical frequencies of altered or reduced expression of p53, PTEN and PAX2, all of which can be appreciated in tubal intraepithelial carcinomas. Because only a subset of these tumors appears to arise in the fallopian tube, attention to expression of these biomarkers in the ovary and other müllerian sites might facilitate the identification of other carcinogenic pathways. PAX2 and PTEN, in addition to p53 and p16(ink4), comprise a potentially important gene combination in high-grade pelvic carcinogenesis.

Endometrial intraepithelial neoplasia.

Endometrial intraepithelial neoplasia (EIN) is a monoclonal premalignant endometrial glandular lesion that precedes the development of endometrioid-type endometrial adenocarcinoma. EIN arises through complex interactions involving the sequential accumulation of genetic damage in endometrial glands and the positive selective pressure of unopposed estrogen. Recent data have revealed a preclinical latent precursor lesion composed of mutated but morphologically nondescript glands that may persist for years in normal-appearing premenopausal cycling endometrium. This latent precursor shares many of the molecular features of EIN and endometrial adenocarcinoma, including frequent inactivation of both the tumor suppressor gene PTEN and the paired box-containing gene PAX2. Upon progression to EIN, the distinctive appearance of crowded and cytologically altered glands heralds a 45-fold increased risk of developing endometrial adenocarcinoma. To preserve the high predictability of EIN for concurrent/ subsequent adenocarcinoma, strict adherence to defined diagnostic criteria is essential. These criteria, as well as potential diagnostic pitfalls and their resolution, are discussed herein.

Endometrial changes from short-term therapy with CDB-4124, a selective progesterone receptor modulator.

Selective progesterone receptor modulators are a class of drugs with progesterone antagonist activity that may confer therapeutic benefit for reproductive disorders in premenopausal women. Endometrial structure, which is dynamically controlled by circulating sex hormones, is likely to be perturbed by progesterone receptor modulators through their progesterone antagonist properties. We examined endometrial histology in 58 premenopausal women treated with the progesterone receptor modulator CDB-4124 (also known as Proellex) for endometriosis or uterine leiomyomata in two clinical trials. Endometrial biopsies obtained after 3 or 6 months with doses of 12.5, 25, or 50 mg daily oral CDB-4124 were reviewed independently by three pathologists. Consensus diagnoses using the World Health Organization hyperplasia scoring system, comments on specific histologic features, and clinical annotation were collected and analyzed. The majority of the endometrial biopsies (103 of 174 biopsies) contained histologic changes that are not seen during normal menstrual cycles. The histology of CDB-4124-treated patients was generally inactive or atrophic, and less frequently, proliferative or secretory, superimposed upon which were novel changes including formation of cystically dilated glands, and secretory changes coexisting with mitoses and apoptotic bodies. With increasing treatment dose and duration, the cysts became predominant and their lining inactive or atrophic. Cystic glands in the CDB-4124-treated subjects correlated with increased endometrial thickness by ultrasound. None of the CDB-4124-treated patients developed endometrial carcinoma or hyperplasia while on therapy. CDB-4124 therapy for 3-6 months produces histologic changes that are sufficiently novel that they might easily be misinterpreted by pathologists, particularly as disordered proliferative or hyperplastic endometrium. Knowledge of the constellation of endometrial changes associated with this agent and other progesterone receptor modulators, including cystic architecture and mixed non-physiologic epithelial changes will prevent misdiagnosis.