High concordance of molecular subtyping between pre-surgical biopsy and surgical resection specimen (matched-pair analysis) in patients with vulvar squamous cell carcinoma using p16- and p53-immunostaining.

OBJECTIVE: Vulvar squamous cell carcinoma (VSCC) can be stratified into three molecular subtypes based on the immunoexpression of p16 and p53: HPV-independent p53-abnormal (p53abn) (most common, biologically aggressive), HPV-associated, with p16-overexpression (second most common, prognostically more favourable) and more recently recognised HPV-independent p53-wildtype (p53wt) (rarest subtype, prognostically intermediate). Our aim was to determine whether molecular subtypes can be reliably identified in pre-operative biopsies and whether these correspond to the subsequent vulvectomy specimen. METHODS: Matched-paired pre-surgical biopsies and subsequent resection specimen of 57 patients with VSCC were analysed for the immunohistochemical expression of p16 and p53 by performing a three-tiered molecular subtyping to test the accuracy rate. RESULTS: Most cases 36/57 (63.2%) belonged to the HPV-independent (p53-abn) molecular subtype, followed by HPV-associated 17/57 (29.8%) and HPV-independent (p53wt) 4/57 (7.0%). The overall accuracy rate on biopsy was 91.2% (52/57): 97.3% for p53-abnormal, 94.1% for p16-overexpression and 50% for p16-neg/p53-wt VSCC. Incorrect interpretation of immunohistochemical p53 staining pattern was the reason for discordant results in molecular subtyping in all five cases. In one case there was an underestimation of p53 pattern (wildtype instead of abnormal/aberrant) and in one case an overestimation of the p53 staining pattern (abnormal/aberrant instead of wildtype). In 3/5 there was a "double positive" staining result (p16 overexpression and abnormal/aberrant p53 staining pattern). In that cases additional molecular workup is required for correct molecular subtyping, resulting in an overall need for molecular examination of 3/57 (3.5%). CONCLUSIONS: Compared to the final resections specimen, the three-tiered molecular classification of VSCC can be determined on pre-surgical biopsies with a high accuracy rate. This enables more precise surgical planning, prediction of the response to (chemo) radiation, selection of targeted therapies and planning of the optimal follow-up strategy for patients in the age of personalised medicine.

Human Papilloma Virus-Independent/p53abnormal Keratinizing Squamous Cell Carcinoma of the Uterine Cervix Associated With Uterine Prolapse.

Knowledge about the morphologic and molecular characteristics of cervical squamous cell carcinomas (CSCCs) associated with uterine prolapse is very limited. Detailed histopathological and immunohistochemical (p16, p53, and cytokeratin 17), as well as molecular evaluation for human papillomavirus (HPV)-DNA and p53-mutational analyses in 4 consecutive CSCCs associated with uterine prolapse with definition of a hitherto not well-described HPV-independent/p53abnormal precursor lesion (HPV-independent cervical intraepithelial neoplasia [CIN; differentiated CIN]) and molecular tumorigenetic pathway. Cases diagnosed within 7 years with a mean age of 75 (range: 69-83) years and a mean tumor size of 7.3 cm (range: 5.2-9.4 cm). All patients presented with locally advanced disease, and 1 woman died of the disease within 4, and another within 14 months of follow-up. All CSCCs and their adjacent precursor lesions were negative for p16, with aberrant p53-expression and diffuse and strong staining for cytokeratin 17. Both the CSCCs and their precursors were negative for HPV-DNA but harbored a TP53 mutation. The precursor lesions were characterized by epithelial thickening with superficial keratinization, and the presence of basal and parabasal keratinocytes with mitotic figures beyond the basal layer, thus showing features similar to those seen in differentiated types of vulvar intraepithelial lesions (vulvar intraepithelial neoplasia [VIN] syn. HPV-independent/p53abn VIN), suggesting the terminology of differentiated CIN or HPV-independent/p53abn CIN. An HPV-independent pathogenetic pathway with a p53-alteration was identified for these cases. CSCC associated with uterine prolapse represents HPV-independent tumors harboring a TP53 mutation. For the first time, a precursor lesion of HPV-independent CSCC of the uterine cervix is described with a differentiated VIN-like morphology, and a separate tumorigenic pathway defined.

Placental pathology in sudden intrauterine death (SIUD) in SARS-CoV-2-positive oligosymptomatic women.

BACKGROUND: Pregnant women are also susceptible to SARS-CoV-2. Although an infection of the placenta may be rare, pregnancy may occasionally be affected by intrauterine failure. The knowledge of placental morphology on sudden intrauterine demise is still limited. METHODS: Fetal and placental tissue of two cases of sudden intrauterine death in the second trimester were analysed morphologically and by immunohistochemistry. One case was evaluated by RT-PCR. RESULTS: Both mothers were tested positive for the Alpha variant of SARS-CoV-2 but were oligosymptomatic for COVID-19. Unexpected sudden intrauterine death (SIUD) occurred at 15 + 2 and 27 + 3 weeks of gestation. One fetus demonstrated an intrauterine growth restriction. No malformations nor inflammatory changes were observed in either fetus on autopsy. In contrast to the placentas, the fetal tissue was negative for SARS-CoV-2 on immunohistochemical and RT-PCR analyses. Macroscopically, the placentas showed an increased consistency with a white, reticular cutting surface covering about 95% of the whole placenta. Only very focal histiocytic chronic intervillositis was noted histologically. Massive perivillous fibrin deposits with extensive necroses of the villous trophoblast were present in more than 90% of the placental tissue. Immunohistochemical staining was strong and diffusely positive for SARS-CoV-2 in the villous trophoblast and rarely within the villous stromal cells. Placental SARS-CoV-2 infection was confirmed by RT-PCR. CONCLUSION: Sudden intrauterine death may occur in mothers who are oligosymptomatic for COVID-19. Acute placental failure is responsible for SIUD, demonstrated by massive perivillous fibrin deposits and extensive necroses of the villous trophoblast with SARS-CoV-2-positivity based on immunohistochemical staining and RT-PCR. Detailed histopathological examination of placental and fetal tissue is mandatory to verify SARS-CoV-2 and to evaluate the pathogenesis and functionality of this disease.

Biallelic pathogenic variants in roundabout guidance receptor 1 associate with syndromic congenital anomalies of the kidney and urinary tract.

Congenital anomalies of the kidney and urinary tract (CAKUT) represent the most common cause of chronic kidney failure in children. Despite growing knowledge of the genetic causes of CAKUT, the majority of cases remain etiologically unsolved. Genetic alterations in roundabout guidance receptor 1 (ROBO1) have been associated with neuronal and cardiac developmental defects in living individuals. Although Slit-Robo signaling is pivotal for kidney development, diagnostic ROBO1 variants have not been reported in viable CAKUT to date. By next-generation-sequencing methods, we identified six unrelated individuals and two non-viable fetuses with biallelic truncating or combined missense and truncating variants in ROBO1. Kidney and genitourinary manifestation included unilateral or bilateral kidney agenesis, vesicoureteral junction obstruction, vesicoureteral reflux, posterior urethral valve, genital malformation, and increased kidney echogenicity. Further clinical characteristics were remarkably heterogeneous, including neurodevelopmental defects, intellectual impairment, cerebral malformations, eye anomalies, and cardiac defects. By in silico analysis, we determined the functional significance of identified missense variants and observed absence of kidney ROBO1 expression in both human and murine mutant tissues. While its expression in multiple tissues may explain heterogeneous organ involvement, variability of the kidney disease suggests gene dosage effects due to a combination of null alleles with mild hypomorphic alleles. Thus, comprehensive genetic analysis in CAKUT should include ROBO1 as a new cause of recessively inherited disease. Hence, in patients with already established ROBO1-associated cardiac or neuronal disorders, screening for kidney involvement is indicated.

Molecular subclassification of vulvar squamous cell carcinoma: reproducibility and prognostic significance of a novel surgical technique.

OBJECTIVES: Vulvar squamous cell carcinoma is subclassified into three prognostically relevant groups: (i) human papillomavirus (HPV) associated, (ii) HPV independent p53 abnormal (mutant pattern), and (iii) HPV independent p53 wild type. Immunohistochemistry for p16 and p53 serve as surrogates for HPV viral integration and TP53 mutational status. We assessed the reproducibility of the subclassification based on p16 and p53 immunohistochemistry and evaluated the prognostic significance of vulvar squamous cell carcinoma molecular subgroups in a patient cohort treated by vulvar field resection surgery. METHODS: In this retrospective cohort study, 68 cases treated by vulvar field resection were identified from the Leipzig School of Radical Pelvic Surgery. Immunohistochemistry for p16 and p53 was performed at three different institutions and evaluated independently by seven pathologists and two trainees. Tumors were classified into one of four groups: HPV associated, HPV independent p53 wild type, HPV independent p53 abnormal, and indeterminate. Selected cases were further interrogated by (HPV RNA in situ hybridization, TP53 sequencing). RESULTS: Final subclassification yielded 22 (32.4%) HPV associated, 41 (60.3%) HPV independent p53 abnormal, and 5 (7.3%) HPV independent p53 wild type tumors. Interobserver agreement (overall Fleiss' kappa statistic) for the four category classification was 0.74. No statistically significant differences in clinical outcomes between HPV associated and HPV independent vulvar squamous cell carcinoma were observed. CONCLUSION: Interobserver reproducibility of vulvar squamous cell carcinoma subclassification based on p16 and p53 immunohistochemistry may support routine use in clinical practice. Vulvar field resection surgery showed no significant difference in clinical outcomes when stratified based on HPV status.

[Practical diagnostic aspects of uterine leiomyosarcoma in the context of the 2020 WHO classification]. / Praktisch-diagnostische Aspekte uteriner Leiomyosarkome im Kontext der WHO-Klassifikation 2020.

The 2020 WHO Classification defines the spindle cell, epithelioid, and myxoid variants as subtypes of uterine leiomyosarcomas (LMS). Presence of cellular atypia (size variation of polymorphic nuclei > 2-3:1), tumor cell necroses, and mitotic count (usually ≥ 10 MF/10 HPF) are still the key features for diagnostic separation from uterine leiomyomas. Preanalytic variables, staining quality, as well as intralesional geographic distribution may affect the mitotic count. Smooth muscle tumors of uncertain malignant potential (STUMP) still exist as a not yet well-characterized diagnostic entity. Immunohistochemical stains against p16, p53, Ki-67, and WT­1 may aid differential diagnosis in selected cases. Diagnostic molecular pathology is not yet relevant for diagnosis.

[Endometrial and other rare uterine sarcomas : Diagnostic aspects in the context of the 2020 WHO classification]. / Endometriale und weitere seltene uterine Sarkome : Diagnostische Aspekte im Kontext der WHO-Klassifikation 2020.

Uterine sarcomas are a heterogeneous group of rare malignancies. Mostly (40-50%), they are leiomyosarcomas, followed by endometrial stromal sarcomas (ESS), low-grade (LG) and high-grade (HG), as well as undifferentiated sarcoma of the uterus (UUS) and adenosarcomas (AS). Other, non-organ-specific tumours such as NTRK-rearranged spindle cell neoplasia, perivascular epithelioid cell tumour (PEComa) and inflammatory myofibroblastic tumour (IMT) are extremely difficult to differentiate.In the most recent WHO classification, endometrial stromal tumours are subdivided as follows: benign, expansively growing endometrial stromal nodule (ESN) with sharp demarcation, the histologically similar-looking LG-ESS with infiltrative growth, the highly malignant HG-ESS and, as a diagnosis of exclusion, the highly aggressive UUS lacking specific lines of differentiation. LG-ESS can be differentiated from HG-ESS in most cases histomorphologically and immunohistochemically, but molecular investigations are necessary in individual cases. HG-ESS can be divided into 4 subtypes (YWHAE/NUTM2 fusion low-grade component, YWHAE/NUTM2 fusion high-grade component, ZC3H7B-BCOR fusion or BCOR-ITD) on the basis of molecular findings. Prognostically unfavourable factors in AS are severe sarcomatous overgrowth, deep myometrial invasion, high-grade histology and lymphatic vessel invasion. Tumours with NTRK fusion are immunohistochemically positive for S100 and TRK. PEComas express cathepsin K and HMB45, as well as TFE3 when translocation is present. Almost every IMT shows an alteration in the ALK gene In the case of overlapping morphology and simultaneous therapeutic and prognostic relevance, it is becoming increasingly important to verify or confirm the suspected histomorphological diagnosis by immunohistochemical and possibly molecular investigations.

[S2k guidelines for the diagnosis and treatment of endometriosis-Recommendations for pathology]. / S2k-Leitlinie Diagnostik und Therapie der Endometriose ­ Anforderungen an die Pathologie.

The present article summarises the recommendations for the handling, histopathological workup, diagnostics and reporting in surgical pathology of biopsies and resection specimens in patients with the clinical diagnosis of endometriosis. In addition to practical aspects of pathology, the guidelines also take into account the clinical requirements for histopathology for the optimal diagnosis and therapy of the patients.Based on the definition of endometriosis of the corpus uteri (adenomyosis uteri) most commonly used in the pathological literature, this was defined in the guidelines as the detection of the endometriosis focus in the myometrium at a distance from the endomyometrial border of a medium-sized visual field (100× magnification), which in metric units corresponds to around 2.5 mm. In bowel resection specimens, the status of the resection margins had to be documented within the histopathological report.Also mentioned are the requirements for the reporting of carcinomas associated with endometriosis, including the immunohistochemical evaluation of steroid hormone receptors and mismatch repair proteins.

[Interdisciplinary S2k guidelines on the diagnosis and treatment of vaginal carcinoma and its precursors-recommendations on surgical pathology for histopathological workup, diagnostics, and reporting]. / S2k-Leitlinie Diagnostik und Therapie des Vaginalkarzinoms und seiner Vorstufen ­ Anforderungen an die Pathologie.

Primary vaginal carcinoma is rare. There are two pathogenetic pathways, one associated with HPV high-risk infection and another one with inactivation of p53. Vaginal Paget's disease is rare and mostly associated with vulvar disease or represents intravaginal spread of associated locoregional cancer. Diagnostic vaginal biopsies should be examined by step sections on H&E. Sentinel lymph nodes should be processed completely using ultrastaging. Morphology-based prognostic factors with good clinical evidence are tumour stage and lymph node status. Molecular markers are not currently relevant for treatment decision and prognosis.

[Current WHO classification of the female genitals : Many new things, but also some old]. / Aktuelle WHO-Klassifikation des weiblichen Genitale : Viel Neues, aber auch manch Altes.

The new WHO classification of tumors of the female genitalia entails some changes, especially those of prognostic and therapeutic relevance: there is a return to the term borderline tumor. Implants are again subdivided into noninvasive implants of the epithelial or desmoplastic type as before. Invasive extraovarian implants are classified as low-grade serous carcinoma (LGSC). Former seromucinous carcinomas are now classified as endometrioid carcinomas (seromucinous subtype). New entities of ovarian carcinomas are mesonephric-like adenocarcinoma, undifferentiated and dedifferentiated carcinoma, and mixed carcinoma. The classification of neuroendocrine neoplasms is analogous to that of pulmonary and gastrointestinal neuroendocrine neoplasms, regardless of their location. Endometrioid endometrial carcinoma can be classified into four molecular subtypes, which have significant prognostic significance. New subtypes include mucinous carcinoma of the intestinal type and mesonephric-like adenocarcinoma. Stromasarcomas of the endometrium are further subclassified based on specific molecular alterations. Adenocarcinomas (ACs) and squamous cell carcinomas (PECs) of the lower female genital tract are distinguished from HPV-associated and HPV-independent carcinomas. Block-like staining for p16 is the accepted surrogate immunohistochemical marker. Grading has not been reported for PEC. For HPV-associated AC of the cervix uteri, prognostic assessment is based on the pattern of invasion (so-called Silva pattern). Serous carcinomas in the cervix uteri are endometrial carcinomas with cervical infiltration.

[Reporting and handling of lymphonodectomy specimens in gynecologic malignancies and sentinel lymph nodes]. / Leitlinienbasierte Aufarbeitung und Befundung von Lymphonodektomiepräparaten und Sentinel-Lymphknoten gynäkologischer Malignome.

The handling and reporting of resected lymph nodes in gynecologic cancer follows the recommendations of the German national guidelines and the recommendations of the International Collaboration of Cancer Reporting (ICCR) and the International Society of Gynecologic Pathologists (ISGyP). The definitions of micrometastases and isolated tumor cells are in accordance with the definition of the UICC (Union Internationale Contre le Cancer) and TNM system. Both findings must be reported as part of the pathology report and final tumor classification. It is mandatory to examine all excised lymph nodes with complete processing of all nodes up to 0.3 cm and slicing of all larger nodes in 0.2-cm wide intervals with complete processing of all lamellae. The amount of the resected lymph nodes in correlation to positive nodes, the metric dimension of the largest lymph node metastasis per lymph node region, and the presence of extracapsular extension of the lymph node deposits must be part of the pathology report. The handling and cutting of sentinel lymph nodes are similar to nonsentinel nodes. Within frozen section analyses and final processing from paraffin-embedded sentinel nodes, all nodes should be examined by three-step sections with an interval of about 200 µm. In cases of negative sentinel nodes on H&E staining, immunohistochemical ultrastaging should be performed.

[Interdisciplinary S2k guidelines on the diagnosis and treatment of uterine sarcomas-recommendations for surgical pathology]. / S2k-Leitlinie Diagnostik und Therapie uteriner Sarkome ­ Anforderungen an die Pathologie.

Uterine sarcomas represent a heterogeneous group of rare malignancies, derived from the myometrium, the endometrial stroma, and very rarely from the nonspecialized uterine soft tissue. The actual incidence is about 1.5 for Caucasian and 3.0 for Afro-American women. There is no grading system for leimoysarcoma defined by the WHO classification; however, if clinicians request, the FNCLCC grading can be specified in analogy to soft tissue sarcomas. Adenosarcomas must be distinguished from adenofibromas (the existence of which is questionable)-with the vast majority of these tumors being uterine adenosarcomas. Within adenosarcomas, deep myometrial invasion (>50%), sarcomatous overgrowth, and a high-grade heterologous component are associated with a higher recurrence rate and poor survival. The immunohistochemical panel represents a very helpful tool for distinguishing low-grade from high grade endometrial stromal sarcomas (ESS) and may be supplemented by molecular analyses. Steroid hormone receptor analysis should be performed for all ESS due to the possible therapeutic relevance. Undifferentiated uterine sarcomas represent a diagnosis of exclusion and have a very poor prognosis. Carcinosarcomas represent a special subtype of endometrial carcinomas and are in fact not uterine sarcomas. Uterine sarcomas may present substantial intratumoral heterogeneity and adequate embedding is mandatory. Lesions ≤2 cm in the largest dimension should be processed completely and larger tumors should be processed with one block per centimeter for the largest tumor dimension.

Histogram Analysis of Diffusion Weighted Imaging at 3T is Useful for Prediction of Lymphatic Metastatic Spread, Proliferative Activity, and Cellularity in Thyroid Cancer.

Pre-surgical diffusion weighted imaging (DWI) is increasingly important in the context of thyroid cancer for identification of the optimal treatment strategy. It has exemplarily been shown that DWI at 3T can distinguish undifferentiated from well-differentiated thyroid carcinoma, which has decisive implications for the magnitude of surgery. This study used DWI histogram analysis of whole tumor apparent diffusion coefficient (ADC) maps. The primary aim was to discriminate thyroid carcinomas which had already gained the capacity to metastasize lymphatically from those not yet being able to spread via the lymphatic system. The secondary aim was to reflect prognostically important tumor-biological features like cellularity and proliferative activity with ADC histogram analysis. Fifteen patients with follicular-cell derived thyroid cancer were enrolled. Lymph node status, extent of infiltration of surrounding tissue, and Ki-67 and p53 expression were assessed in these patients. DWI was obtained in a 3T system using b values of 0, 400, and 800 s/mm². Whole tumor ADC volumes were analyzed using a histogram-based approach. Several ADC parameters showed significant correlations with immunohistopathological parameters. Most importantly, ADC histogram skewness and ADC histogram kurtosis were able to differentiate between nodal negative and nodal positive thyroid carcinoma. CONCLUSIONS: histogram analysis of whole ADC tumor volumes has the potential to provide valuable information on tumor biology in thyroid carcinoma. However, further studies are warranted.

MRI Texture Analysis of Inguinal Lymph Node Metastasis in Vulvar Cancer - Associations With Histopathology.

BACKGROUND/AIM: Texture analysis is a quantitative imaging technique that provides novel biomarkers beyond conventional image reading. This study aimed to investigate the correlation between texture parameters and histopathological features of lymph nodes in patients with vulvar cancer. PATIENTS AND METHODS: Overall, nine female patients (mean age 70.1±13.4 years, range=39-87 years) were included in the analysis. All patients had squamous cell carcinomas and underwent upfront surgery with inguinal lymph node resection. Immunohistochemical assessment was performed using several markers of the epithelial-mesenchymal transition. The presurgical magnetic resonance imaging (MRI) was analyzed with the MaZda package. RESULTS: In discrimination analysis, several parameters derived from T1-weighted images showed statistically significant differences between non-metastatic and metastatic lymph nodes. The highest statistical significance was reached by the texture feature "S(0,3)InvDfMom" (p=0.016). In correlation analysis, significant associations were found between MRI texture parameters derived from both T1-weighted and T2-weighted images and the investigated histopathological features. Notably, S(0,3)InvDfMom derived from T1-weighted images highly correlated with the Vimentin-score (r=0.908, p=0.001). CONCLUSION: Several associations between MRI texture analysis and immunohistochemical parameters were identified in metastasized lymph nodes of cases with vulvar cancer.

Diagnostic accuracy and reliability of CT-based Node-RADS for colon cancer.

OBJECTIVE: The Node-RADS classification was recently published as a classification system to better characterize lymph nodes in oncological imaging. The present analysis investigated the diagnostic benefit of the Node-RADS classification of staging computed tomography (CT) images to categorize and stage lymph nodes in patients with colon cancer. MATERIALS AND METHODS: All patients were surgically resected and the lymph nodes were histopathological analyzed. All investigated lymph nodes were scored in accordance to the Node-RADS classification by two experienced radiologists. Interreader variability was assessed with Cohen's kappa analysis, discrimination analysis was performed with Mann-Whitney-U test and diagnostic accuracy was assessed with receiver-operating characteristics (ROC) curve analysis. RESULTS: Overall, 108 patients (n = 49 females, 45.3%) with a mean age of 70.08 ± 14.34 years were included. In discrimination analysis, the total Node-RADS score showed statistically significant differences between N- and N + stage (for reader 1: mean 1.89 ± 1.09 score for N- versus 2.93 ± 1.62 score for N+, for reader 2: 1.33 ± 0.48 score for N- versus 3.65 ± 0.94 score for N+, p = 0.001, respectively). ROC curve analysis for lymph node discrimination showed an area under the curve of 0.68. A threshold value of 2 resulted in a sensitivity of 0.62 and a specificity of 0.71. CONCLUSION: Node-RADS score derived from staging CT shows only limited diagnostic accuracy to correctly predict nodal positivity in colon cancer. The interreader variability seems to be high and should question the clinical translation for this tumour entity.

Impact of CT texture analysis on complication rate in CT-guided liver biopsies.

Aim of the study: Texture analysis derived from computed tomography (CT) involves quantitative imaging parameters characterizing possible valuable associations with clinical purposes. Their prognostic capability in patients undergoing percutaneous CT-guided liver biopsy to identify associations with postinterventional bleeding complications and biopsy success is not sufficiently explored. Material and methods: Three hundred fifteen patients (124 female, 39%) with a mean age of 62.5 ±10.2 years underwent percutaneous CT-guided liver biopsy and were analyzed regarding clinical, procedure-related, and CT texture features. Results: Thirty patients (9.5%) presented with bleeding after biopsy (including two requiring interventional treatment), whereas 46 patients (14.6%) had negative biopsy successes. Distance of lesion from liver capsule was statistically significantly different in patients with and without bleeding (p = 0.015). Several texture features were statistically significantly different between the groups, S(0,1)SumAverg having the highest significance (p = 0.004). Regarding unsuccessful biopsy results, liver fibrosis was the only clinical feature with statistical significance (p = 0.049). Only two texture features (S(4,-4)InvDfMom and Teta3) were statistically different between the groups according to the biopsy result. Conclusions: Several CT texture features of the target lesion and the length from the capsule to the lesion were associated with bleeding complications after CT-guided percutaneous liver biopsy. This could be used to identify patients at risk at the beginning of the procedure.

Mucinous cystadenoma and benign mesonephric-like proliferation in the ovary - Further evidence for clonal relationship.

Mesonephric-like adenocarcinomas rarely occur in the uterus and the ovary. Benign mesonephric-like (ML) proliferations and hyperplasia have been described solely within the ovary. Pathogenetic data are very limited. We report a case with microscopic focus of benign ML-proliferation in association with mucinous cystadenoma in the ovary. The immunophenotype was distinct (mucinous tumor: focal weak nuclear positivity for PAX-8, CK 7, patchy cytoplasmic positivity for p16 and negativity for estrogen receptor, CD 10, TTF-1, p53 wildtype; mesonephric component: diffusely positive for PAX-8, CK 7, luminal CD 10, TTF-1, focal staining for estrogen receptor, patchy cytoplasmic for p16, p53 wildtype). On NGS-analysis there was clonal mutation of KRAS p.G12C. The data provide additional evidence for the concept of transdifferentiation (Müllerian tissue representing Wolffian/mesonephric features on histology and immunostaining) within the pathogenesis of mesonephric proliferation of the female genital tract and demonstrate the clonal relationship between these distinct morphologic components.

Multi-Parameter Analysis of Disseminated Tumor Cells (DTCs) in Early Breast Cancer Patients with Hormone-Receptor-Positive Tumors.

BACKGROUND: Patients with hormone-receptor-positive (HR+) breast cancer are at increased risk for late recurrence. One reason might be disseminated tumor cells (DTCs), which split off in the early stages of the disease and metastasize into the bone marrow (BM). METHODS: We developed a novel multi-parameter immunofluorescence staining protocol using releasable and bleachable antibody-fluorochrome-conjugates. This sequential procedure enabled us to analyze six distinct phenotypical and therapy-related markers on the same DTC. We characterized BM aspirates from 29 patients with a HR+ tumor and a known positive DTC status-based on the standardized detection of epithelial cells in BM. RESULTS: Using the immunofluorescence staining, a total of 153 DTCs were detected. Luminal A patients revealed a higher DTC count compared with luminal B. The majority of the detected DTCs were CK-positive (128/153). However, in 16 of 17 luminal A patients we found HER2-positive DTCs. We detected CK-negative DTCs (25/153) in 12 of 29 patients. Of those cells, 76% were Ki67-positive and 68% were HER2-positive. Moreover, we detected DTC clusters consisting of mixed characteristics in 6 of 29 patients. CONCLUSIONS: Using sequential multi-parameter imaging made it possible to identify distinct DTC profiles not solely based on epithelial features. Our findings indicate that characterization rather than quantification of DTCs might be relevant for treatment decisions.

[Molecular classification of endometrial carcinoma-a short summary for clinical use]. / Molekulare Klassifikation des Endometriumkarzinoms ­ ein kurzer Überblick.

BACKGROUND: Histopathological examination is still the backbone for the diagnosis and treatment decision making in endometrial carcinoma (EC). The binary classification of EC into type 1 (mostly endometrioid) and type 2 (mostly serous), although still helpful, showed overlapping clinical, morphological and molecular features and was not very prognostic discriminatory for all subtypes of EC. METHODS: Analysing the most recent studies dealing with the molecular classification of EC and the recommendations of the German S3-guidelines for EC. RESULTS AND CONCLUSION: Based on the comprehensive molecular study of The Cancer Genome Atlas Project (TCGA) four distinct molecular subtypes have been identified: EC with POLE mutation (POLEmut), with loss of mismatch repair proteins (MMR deficiency; dMMR), or with TP53 mutation (p53mut) and without any of these alterations, termed NSMP (no specific molecular profile). The molecular classification of EC presents a morphomolecular approach, based on histopathological evaluation (tumor diagnosis, subtyping, grading), immunohistochemistry (MMR, p53) and molecular analyses for POLE. The incorporation of this molecular classification is recommended for clinical use by the World Health Organisation (WHO) as well as many national guidelines and international societies. Due to the heterogeneity of NSMP-EC, which is the largest molecular group, additional research is indicated to further characterise these tumors.

Mesonephric-like adenocarcinoma of the female genital tract: possible role of KRAS-targeted treatment-detailed molecular analysis of a case series and review of the literature for targetable somatic KRAS-mutations.

PURPOSE: Mesonephric-like adenocarcinomas (MLA) of the female genital tract represent a rare and relatively recently described neoplasm exhibiting characteristic morphologic and immunohistochemical findings commonly associated with a KRAS-mutation. Most cases display an aggressive clinical behavior, but knowledge about treatment approaches is limited, especially for targeting KRAS. METHODS: We report a series of eight cases with a detailed molecular analysis for KRAS. These cases as well as the data of previously published cases with detailed information regarding KRAS-mutational events were reviewed for a potential targeted approach and its prognostic impact. RESULTS: Both the uterine and ovarian MLA harbor a somatic KRAS-mutation in about 85% of the reported cases, affecting the hotspot codons 12 and 13. 15.7% of the endometrial and 15.6% of ovarian MLA are wild type for KRAS. A p.G12A-alteration was seen in 5.6% (5/89) of the endometrial and in 6.2% (2/32) of the ovarian tumors, for p.G12C in 7.9% and 6.2%, for p.G12D in 32.6% and 34.5% and for p.G12V in 36% and 37.5%, respectively. Very limited data are available regarding the prognostic impact of different mutational sites within the KRAS-gene without significant prognostic impact. CONCLUSION: Because of a specific p.G12C-KRAS somatic mutation, only the minority of MLA (7.9% with uterine and 6.2% with ovarian primary) are potentially targetable by sotarasib in that rare but aggressive subtype of adenocarcinoma of the female genital tract. Until now, the different location of a somatic KRAS-mutation is of no prognostic impact.