High-fat diet and obesity are associated with differential angiogenic gene expression in epithelial ovarian cancer.

OBJECTIVE: We sought to evaluate the association between diet and angiogenic biomarkers in KpB mice, and the association between these markers, body mass index (BMI), and overall survival (OS) in high-grade serous cancers (HGSC). METHODS: Tumors previously obtained from KpB mice subjected to high-fat diets (HFD, n = 10) or low-fat diets (LFD, n = 10) were evaluated for angiogenesis based on CD-31 microvessel density (MVD). Data from prior microarray analysis (Agilent 244 K arrays) conducted in 10 mice were utilized to assess associations between diet and angiogenetic biomarkers. Agilent (mouse) and Affymetrix Human Genome U133a probes were linked to 162 angiogenic-related genes. The associations between biomarkers, BMI, and OS were evaluated in an HGSC internal database (IDB) (n = 40). Genes with unadjusted p < 0.05 were evaluated for association with OS in the TCGA-OV database (n = 339). RESULTS: There was no association between CD-31 and diet in mice (p = 0.66). Sixteen angiogenic-related genes passed the p < 0.05 threshold for association with HFD vs. LFD. Transforming growth factor-alpha (TGFA) demonstrated 72% higher expression in HFD vs. LFD mice (p = 0.04). Similar to the mouse study, in our HGSC IDB, higher TGFA expression correlated with higher BMI (p = 0.01) and shorter survival (p = 0.001). In the TCGA-OV dataset, BMI data was not available and there was no association between TGFA and OS (p = 0.48). CONCLUSIONS: HFD and obesity may promote tumor progression via differential modulation of TGFA. We were unable to confirm this finding in the TCGA dataset. Further evaluation of TGFA is needed to determine if this is a target unique to obesity-driven HGSC.

EWSR1::NR4A3 gene fusion in a cutaneous atypical myoepithelial neoplasm.

Myoepithelial neoplasms of the skin and soft tissue are rare and share histopathologic features with their salivary gland counterpart. We present a case of an atypical myoepithelial neoplasm from the back of a 72-year-old female. This lesion harbored an EWSR1::NR4A3 gene fusion, a genetic signature characteristically seen in extraskeletal myxoid chondrosarcoma. To our knowledge, this is a unique case of an atypical cutaneous myoepithelial neoplasm harboring EWSR1::NR4A3 fusion.

BAP1 Tumor Predisposition Syndrome Presenting as a Recurrent Ovarian Sex Cord-Stromal Tumor.

The BRCA1-associated protein 1 ( BAP1 ) gene encodes a tumor suppressor that functions as a ubiquitin hydrolase involved in DNA damage repair. BAP1 germline mutations are associated with increased risk of multiple solid malignancies, including mesothelioma, uveal melanoma, renal cell carcinoma, and high-grade rhabdoid meningiomas. Here, we describe the case of a 52-yr-old woman who experienced multiple abdominal recurrences of an ovarian sex cord-stromal tumor that was originally diagnosed at age 25 and who was found to have a germline mutation in BAP1 and a family history consistent with BAP1 tumor predisposition syndrome. Recurrence of the sex cord-stromal tumor demonstrated loss of BAP1 expression by immunohistochemistry. Although ovarian sex cord-stromal tumors have been described in mouse models of BAP1 tumor predisposition syndrome, this relationship has not been previously described in humans and warrants further investigation. The case presentation, tumor morphology, and immunohistochemical findings have overlapping characteristics with peritoneal mesotheliomas, and this case represents a potential pitfall for surgical pathologists.

Giant Pancreatic Myoepithelial Hamartoma in a Patient With Familial Insulin-Dependent Diabetes Mellitus.

ABSTRACT: Pancreatic myoepithelial hamartoma is a rare, benign solid and cystic lesion of the pancreas. We present the first case of an adult with a giant myoepithelial hamartoma extending throughout the pancreas in a patient with diabetes in 4 immediate family members. The patient is a 46-year-old man presented with recurrent acute pancreatitis. Computed tomographic imaging showed that the head and body of the pancreas were replaced by a solid-cystic mass with focal calcification. Medical history includes insulin-dependent diabetes mellitus (IDDM) diagnosed at age 30. Endoscopic ultrasound-guided fine-needle aspiration showed pancreatic acinar tissue and smooth muscle without evidence of malignancy. Total pancreatectomy was performed because of the diffuse nature of the cystic disease and preexisting IDDM. The histopathologic diagnosis was consistent with myoepithelial hamartoma. In addition, there was a family history of IDDM and hamartomatous cyst resection in the paternal grandmother. We report the first case of diffuse pancreatic myoepithelial hamartoma with near total replacement of the entire pancreatic parenchyma, and the first reported case associated with a family history of heritable IDDM. Improved knowledge of the genetics, development, and malignant potential of such rare diseases is critical to determine appropriate management for patients.

Identification and targeting of a HES1-YAP1-CDKN1C functional interaction in fusion-negative rhabdomyosarcoma.

Rhabdomyosarcoma (RMS), a cancer characterized by features of skeletal muscle, is the most common soft-tissue sarcoma of childhood. With 5-year survival rates among high-risk groups at &lt; 30%, new therapeutics are desperately needed. Previously, using a myoblast-based model of fusion-negative RMS (FN-RMS), we found that expression of the Hippo pathway effector transcriptional coactivator YAP1 (YAP1) permitted senescence bypass and subsequent transformation to malignant cells, mimicking FN-RMS. We also found that YAP1 engages in a positive feedback loop with Notch signaling to promote FN-RMS tumorigenesis. However, we could not identify an immediate downstream impact of this Hippo-Notch relationship. Here, we identify a HES1-YAP1-CDKN1C functional interaction, and show that knockdown of the Notch effector HES1 (Hes family BHLH transcription factor 1) impairs growth of multiple FN-RMS cell lines, with knockdown resulting in decreased YAP1 and increased CDKN1C expression. In silico mining of published proteomic and transcriptomic profiles of human RMS patient-derived xenografts revealed the same pattern of HES1-YAP1-CDKN1C expression. Treatment of FN-RMS cells in vitro with the recently described HES1 small-molecule inhibitor, JI130, limited FN-RMS cell growth. Inhibition of HES1 in vivo via conditional expression of a HES1-directed shRNA or JI130 dosing impaired FN-RMS tumor xenograft growth. Lastly, targeted transcriptomic profiling of FN-RMS xenografts in the context of HES1 suppression identified associations between HES1 and RAS-MAPK signaling. In summary, these in vitro and in vivo preclinical studies support the further investigation of HES1 as a therapeutic target in FN-RMS.

Extended Human Papillomavirus Genotyping to Predict Progression to High-Grade Cervical Precancer: A Prospective Cohort Study in the Southeastern United States.

BACKGROUND: High-risk human papillomavirus (hrHPV) testing is utilized in primary cervical cancer screening, generally along with cytology, to triage abnormalities to colposcopy. Most screening-based hrHPV testing involves pooled detection of any hrHPV or of HPV16/18. Cervical neoplasia progression risks based on extended hrHPV genotyping-particularly non-16/18 hrHPV types-are not well characterized. HPV genotype-specific incidence of high-grade cervical intraepithelial neoplasia or more severe (CIN2+) following an abnormal screening result was examined. METHODS: We assessed a US-based prospective, multiracial, clinical cohort of 343 colposcopy patients with normal histology (n = 226) or CIN1 (n = 117). Baseline cervical samples underwent HPV DNA genotyping, and participants were followed up to 5 years. Genotype-specific CIN2+ incidence rates (IR) were estimated with accelerated failure time models. Five-year CIN2+ risks were estimated nonparametrically for hierarchical hrHPV risk groups (HPV16; else HPV18/45; else HPV31/33/35/52/58; else HPV39/51/56/59/68). RESULTS: At enrollment, median participant age was 30.1 years; most (63%) were hrHPV-positive. Over follow-up, 24 participants progressed to CIN2+ (7.0%). CIN2+ IR among hrHPV-positive participants was 3.4/1,000 person-months. CIN2+ IRs were highest for HPV16 (8.3), HPV33 (7.8), and HPV58 (4.9). Five-year CIN2+ risk was higher for HPV16 (0.34) compared with HPV18/45 (0.12), HPV31/33/35/52/58 (0.12), and HPV39/51/56/59/68 (0.16) (P = 0.05). CONCLUSIONS: Non-16/18 hrHPV types are associated with differential CIN2+ progression rates. HPV16, 33, and 58 exhibited the highest rates over 5 years. HPV risk groups warrant further investigation in diverse US populations. IMPACT: These novel data assessing extended HPV genotyping in a diverse clinical cohort can inform future directions to improve screening practices in the general population.

Communicating Certainty in Pathology Reports.

CONTEXT.­: Pathology reports are the main modality in which results are communicated to other physicians. For various reasons, the diagnosis may be qualified on a spectrum of uncertainty. OBJECTIVE.­: To examine how communication of uncertainty is an unexamined source of possible medical error. No study to our knowledge has examined pathology reports across multiple institutions. This study seeks to identify commonly used phrases of diagnostic uncertainty and their interpreted meanings by surgical pathologists and clinicians. DESIGN.­: Anonymous surveys were completed at 3 major US academic institutions by 18 practicing staff pathologists, 12 pathology residents, 53 staff clinicians, and 50 resident/allied health professional clinicians at 5 standard tumor boards. All participants rated percentage certainty associated with 7 diagnostic terms. Pathologists answered 2 questions related to the ability to clarify a diagnosis using a comment and comfort wording pathology reports. Clinicians answered questions on how often they read a pathology report comment, if they found the comment helpful, and how comfortable they were in reading pathology reports. RESULTS.­: A wide range in percentage certainty was found for each of the 7 diagnostic phrases. Both staff and resident clinicians and residents showed wide variability in interpreting the phrases. Twenty-five of 50 staff clinicians (52%) were very comfortable reading a pathology report, whereas only 4 of 53 resident clinicians (8%) were very comfortable reading a pathology report. Twenty-four of 53 staff clinicians (63%) reported always reading the comment, yet only 20 of 53 (27%) always found the comment helpful. The phrases "diagnostic of" and "consistent with" had the strongest agreement in meaning. The weakest agreement was between "suspicious for" and "compatible with." CONCLUSIONS.­: Efforts to standardize diagnostic terms may improve communication.

A Hybrid Human-Machine Learning Approach for Screening Prostate Biopsies Can Improve Clinical Efficiency Without Compromising Diagnostic Accuracy.

CONTEXT.­: Prostate cancer is a common malignancy, and accurate diagnosis typically requires histologic review of multiple prostate core biopsies per patient. As pathology volumes and complexity increase, new tools to improve the efficiency of everyday practice are keenly needed. Deep learning has shown promise in pathology diagnostics, but most studies silo the efforts of pathologists from the application of deep learning algorithms. Very few hybrid pathologist-deep learning approaches have been explored, and these typically require complete review of histologic slides by both the pathologist and the deep learning system. OBJECTIVE.­: To develop a novel and efficient hybrid human-machine learning approach to screen prostate biopsies. DESIGN.­: We developed an algorithm to determine the 20 regions of interest with the highest probability of malignancy for each prostate biopsy; presenting these regions to a pathologist for manual screening limited the initial review by a pathologist to approximately 2% of the tissue area of each sample. We evaluated this approach by using 100 biopsies (29 malignant, 60 benign, 11 other) that were reviewed by 4 pathologists (3 urologic pathologists, 1 general pathologist) using a custom-designed graphical user interface. RESULTS.­: Malignant biopsies were correctly identified as needing comprehensive review with high sensitivity (mean, 99.2% among all pathologists); conversely, most benign prostate biopsies (mean, 72.1%) were correctly identified as needing no further review. CONCLUSIONS.­: This novel hybrid system has the potential to efficiently triage out most benign prostate core biopsies, conserving time for the pathologist to dedicate to detailed evaluation of malignant biopsies.

Association between PEG3 DNA methylation and high-grade cervical intraepithelial neoplasia.

BACKGROUND: Epigenetic mechanisms are hypothesized to contribute substantially to the progression of cervical intraepithelial neoplasia (CIN) to cervical cancer, although empirical data are limited. METHODS: Women (n = 419) were enrolled at colposcopic evaluation at Duke Medical Center in Durham, North Carolina. Human papillomavirus (HPV) was genotyped by HPV linear array and CIN grade was ascertained by biopsy pathologic review. DNA methylation was measured at differentially methylated regions (DMRs) regulating genomic imprinting of the IGF2/H19, IGF2AS, MESTIT1/MEST, MEG3, PLAGL1/HYMAI, KvDMR and PEG10, PEG3 imprinted domains, using Sequenom-EpiTYPER assays. Logistic regression models were used to evaluate the associations between HPV infection, DMR methylation and CIN risk overall and by race. RESULTS: Of the 419 participants, 20 had CIN3+, 52 had CIN2, and 347 had ≤ CIN1 (CIN1 and negative histology). The median participant age was 28.6 (IQR:11.6) and 40% were African American. Overall, we found no statistically significant association between altered methylation in selected DMRs and CIN2+ compared to ≤CIN1. Similarly, there was no significant association between DMR methylation and CIN3+ compared to ≤CIN2. Restricting the outcome to CIN2+ cases that were HR-HPV positive and p16 staining positive, we found a significant association with PEG3 DMR methylation (OR: 1.56 95% CI: 1.03-2.36). CONCLUSIONS: While the small number of high-grade CIN cases limit inferences, our findings suggest an association between altered DNA methylation at regulatory regions of PEG3 and high grade CIN in high-risk HPV positive cases.

Limited Reporting of Histopathologic Details in a Multi-Institutional Academic Cohort of Phyllodes Tumors: Time for Standardization.

BACKGROUND: Phyllodes tumors are rare fibroepithelial neoplasms that are classified by tiered histopathologic features. While there are protocols for the reporting of cancer specimens, no standardized reporting protocol exists for phyllodes. METHODS: We performed an 11-institution contemporary review of phyllodes tumors. Granular histopathologic details were recorded, including the features specifically considered for phyllodes grade classification. RESULTS: Of 550 patients, median tumor size was 3.0 cm, 68.9% (n = 379) of tumors were benign, 19.6% (n = 108) were borderline, and 10.5% (n = 58) were malignant. All cases reported the final tumor size and grade classification. Complete pathologic reporting of all histopathologic features was present in 15.3% (n = 84) of cases, while an additional 35.6% (n = 196) were missing only one or two features in the report. Individual details regarding the degree of stromal cellularity was not reported in 53.5% (n = 294) of cases, degree of stromal atypia in 58.0% (n = 319) of cases, presence of stromal overgrowth in 56.2% (n = 309) of cases, stromal cell mitoses in 37.5% (n = 206) of cases, and tumor border in 54.2% (n = 298) of cases. The final margin status (negative vs. positive) was omitted in only 0.9% of cases, and the final negative margin width was specifically reported in 73.8% of cases. Reporting of details was similar across all sites. CONCLUSION: In this academic cohort of phyllodes tumors, one or more histopathologic features were frequently omitted from the pathology report. While all features were considered by the pathologist for grading, this limited reporting reflects a lack of reporting consensus. We recommend that standardized reporting in the form of a synoptic-style cancer protocol be implemented for phyllodes tumors, similar to other rare tumors.

Endometrial Adenocarcinomas With No Specific Molecular Profile: Morphologic Features and Molecular Alterations of "Copy-number Low" Tumors.

The study evaluated morphologic patterns, mutational profiles, and ß-catenin immunohistochemistry (IHC) in copy-number low (CNL) endometrial adenocarcinomas (EAs). CNL EAs (n=19) with next-generation or whole genome sequencing results and available tissue for IHC were identified from our institutional database. Clinical data and histologic slides were reviewed. IHC for ß-catenin was performed and correlated with mutation status. Images of digital slides of CNL EAs from The Cancer Genome Atlas (TCGA) database (n=90) were blindly reviewed by 4 pathologists, and morphology was correlated with mutation status. Categorical variables were analyzed using the Fisher exact test, and agreement was assessed using Fleiss κ. CTNNB1 mutations were present in 63% (12/19) of CNL EAs. ß-catenin nuclear localization was present in 83% of CTNNB1-mutated tumors (10/12) and in 0% (0/7) of CTNNB1-wildtype tumors (sensitivity 0.83, specificity 1.00). Squamous differentiation (SD) was present in 47% (9/19) and was more often observed in CTNNB1-mutated tumors (P=0.02). Mucinous differentiation (MD) was associated with KRAS mutations (P<0.01). Digital image review of TCGA CNL EAs revealed that pathologist agreement on SD was strong (κ=0.82), whereas agreement on MD was weak (κ=0.48). Pathologists identified SD in 22% (20/90), which was significantly associated with the presence of CTNNB1 mutations (P<0.01). CNL EAs demonstrate several morphologies with divergent molecular profiles. SD was significantly associated with CTNNB1 mutations and nuclear localization of ß-catenin in these tumors. Nuclear expression of ß-catenin is a sensitive and specific IHC marker for CTNNB1 mutations in CNL EAs. CNL EAs with KRAS mutations often displayed MD.

More Than Meets the Eye? A Cautionary Tale of Malignant Ectomesenchymoma Treated as Low-risk Orbital Rhabdomyosarcoma.

Malignant ectomesenchymoma (MEM) is a rare multiphenotypic tumor comprised of mesenchymal and neuroectodermal components. MEM is typically diagnosed in infants and younger children and outcomes are variable. The current approach for treating MEM includes targeting the more aggressive mesenchymal component of the tumor, which is often rhabdomyosarcoma. Here, we describe a case of an orbital tumor initially diagnosed and treated as low-risk rhabdomyosarcoma. Local failure prompting a second biopsy revealed neuronal differentiation consistent with a diagnosis of MEM. Intensifying therapy and local radiotherapy led to a long-term cure. This case offers a cautionary tale that while outcomes for MEM were similar to matched rhabdomyosarcoma cohorts when treated on conventional Intergroup Rhabdomyosarcoma Study Group (IRSG) III/IV protocols, treating MEM using a decreased intensity low-risk rhabdomyosarcoma regimen may not be sufficient.

Epithelioid Hyalinizing Sarcoma With MGA-NUTM1 Fusion.

OBJECTIVES: Soft tissue sarcomas are a group of tumors derived from the mesenchymal origin. Historically, they have been classified according to morphologic and immunohistochemical characteristics. The advent of multiplexed next-generation sequencing (NGS), specifically RNA sequencing, has modified the classification of such tumors and others by determining categorization based on molecular alterations. The NUTM1 rearrangement, previously thought to be present only in carcinomas, has recently been reported in poorly differentiated high-grade sarcomas of the soft tissue. We present the first reported case of an epithelioid hyalinizing sarcoma harboring the MGA-NUTM1 fusion in an acral site. METHODS: Histopathologic, immunohistochemical, and molecular testing were performed on resection tissue. RESULTS: Histologically, the tumor showed an epithelioid morphology with prominent background hyalinization. Immunohistochemically, the tumor expressed CD99 and nuclear NUT-1. By NGS the tumor harbors MGA-NUTM1 fusion. CONCLUSIONS: Our findings support more extensive use of NGS for accurate sarcoma classification and identification of potential therapeutic targets. Furthermore, they corroborate the fact that NUTM1-rearranged soft tissue tumors represent a spectrum of heterogeneous morphologic entities. This case also highlights the utility of NUT-1 immunohistochemical study as a possible screening tool for NUTM1-fused sarcomas.

Development and Validation of the Gene Expression Predictor of High-grade Serous Ovarian Carcinoma Molecular SubTYPE (PrOTYPE).

PURPOSE: Gene expression-based molecular subtypes of high-grade serous tubo-ovarian cancer (HGSOC), demonstrated across multiple studies, may provide improved stratification for molecularly targeted trials. However, evaluation of clinical utility has been hindered by nonstandardized methods, which are not applicable in a clinical setting. We sought to generate a clinical grade minimal gene set assay for classification of individual tumor specimens into HGSOC subtypes and confirm previously reported subtype-associated features. EXPERIMENTAL DESIGN: Adopting two independent approaches, we derived and internally validated algorithms for subtype prediction using published gene expression data from 1,650 tumors. We applied resulting models to NanoString data on 3,829 HGSOCs from the Ovarian Tumor Tissue Analysis consortium. We further developed, confirmed, and validated a reduced, minimal gene set predictor, with methods suitable for a single-patient setting. RESULTS: Gene expression data were used to derive the predictor of high-grade serous ovarian carcinoma molecular subtype (PrOTYPE) assay. We established a de facto standard as a consensus of two parallel approaches. PrOTYPE subtypes are significantly associated with age, stage, residual disease, tumor-infiltrating lymphocytes, and outcome. The locked-down clinical grade PrOTYPE test includes a model with 55 genes that predicted gene expression subtype with >95% accuracy that was maintained in all analytic and biological validations. CONCLUSIONS: We validated the PrOTYPE assay following the Institute of Medicine guidelines for the development of omics-based tests. This fully defined and locked-down clinical grade assay will enable trial design with molecular subtype stratification and allow for objective assessment of the predictive value of HGSOC molecular subtypes in precision medicine applications.See related commentary by McMullen et al., p. 5271.

Frequent Mutations of POT1 Distinguish Pulmonary Sarcomatoid Carcinoma From Other Lung Cancer Histologies.

INTRODUCTION: Pulmonary sarcomatoid carcinoma (PSC) is a rare subtype of non-small-cell lung cancer (NSCLC) harboring mutations in many canonical NSCLC-driver genes (eg, TP53, KRAS, MET). Protection of telomeres 1 (POT1) mutations are observed in angiosarcoma and chronic lymphocytic leukemia, but their frequency in other solid tumors, including NSCLC subtypes, has not been rigorously explored. MATERIALS AND METHODS: We analyzed next-generation sequencing data from 62,368 tumors, including 11,134 NSCLCs and 100 PSCs. We performed logistic regression to identify associations between POT1 mutation frequency and tumor histology across 184 tumor categories, adjusting for tumor mutational burden. We further explored co-occurring gene mutations in genes previously reported to underlie PSC tumorigenesis. RESULTS: Across 184 tumor categories, POT1 mutations were most frequent in PSC and were 14 times more common in PSC (28%) than in other tumor types (P = 1.23 × 10-31) and 6.7 times more common in PSC than other NSCLCs (P = 5.1 × 10-17). PSCs harboring KRAS mutations were significantly more likely to harbor POT1 mutations (P = 1.3 × 10-3), whereas those with TP53 mutations were less likely to harbor POT1 mutations (P = .037). One-fourth of POT1-mutated PSCs harbored a second POT1 mutation. Across all PSCs, 83% of POT1 mutations were in the OB1/OB2 (DNA-binding) domain (P = 1.5 × 10-5), an enrichment not observed in other tumor types. CONCLUSION: We report an unanticipated association between POT1 mutation and PSC. Unlike other molecular alterations that are frequent across NSCLC subtypes, POT1 mutations are largely unique to PSC. This finding may help to develop disease-defining molecular subgroups within PSC and presents opportunities for molecularly stratified prognostication and therapy.

POT1 mutation spectrum in tumour types commonly diagnosed among POT1-associated hereditary cancer syndrome families.

BACKGROUND: The shelterin complex is composed of six proteins that protect and regulate telomere length, including protection of telomeres 1 (POT1). Germline POT1 mutations are associated with an autosomal dominant familial cancer syndrome presenting with diverse malignancies, including glioma, angiosarcoma, colorectal cancer and melanoma. Although somatic POT1 mutations promote telomere elongation and genome instability in chronic lymphocytic leukaemia, the contribution of POT1 mutations to development of other sporadic cancers is largely unexplored. METHODS: We performed logistic regression, adjusted for tumour mutational burden, to identify associations between POT1 mutation frequency and tumour type in 62 368 tumours undergoing next-generation sequencing. RESULTS: A total of 1834 tumours harboured a non-benign mutation of POT1 (2.94%), of which 128 harboured a mutation previously reported to confer familial cancer risk in the setting of germline POT1 deficiency. Angiosarcoma was 11 times more likely than other tumours to harbour a POT1 mutation (p=1.4×10-20), and 65% of POT1-mutated angiosarcoma had >1 mutations in POT1. Malignant gliomas were 1.7 times less likely to harbour a POT1 mutation (p=1.2×10-3) than other tumour types. Colorectal cancer was 1.2 times less likely to harbour a POT1 mutation (p=0.012), while melanoma showed no differences in POT1 mutation frequency versus other tumours (p=0.67). CONCLUSIONS: These results confirm a role for shelterin dysfunction in angiosarcoma development but suggest that gliomas arising in the context of germline POT1 deficiency activate a telomere-lengthening mechanism that is uncommon in gliomagenesis.

Loss of MST/Hippo Signaling in a Genetically Engineered Mouse Model of Fusion-Positive Rhabdomyosarcoma Accelerates Tumorigenesis.

A hallmark of fusion-positive alveolar rhabdomyosarcoma (aRMS) is the presence of a chromosomal translocation encoding the PAX3-FOXO1 fusion oncogene. Primary cell-based modeling experiments have shown that PAX3-FOXO1 is necessary, but not sufficient for aRMS tumorigenesis, indicating additional molecular alterations are required to initiate and sustain tumor growth. Previously, we showed that PAX3-FOXO1-positive aRMS is promoted by dysregulated Hippo pathway signaling, as demonstrated by increased YAP1 expression and decreased MST activity. We hypothesized that ablating MST/Hippo signaling in a genetically engineered mouse model (GEMM) of aRMS would accelerate tumorigenesis. To this end, MST1/2-floxed (Stk3F/F;Stk4F/F ) mice were crossed with a previously established aRMS GEMM driven by conditional expression of Pax3:Foxo1 from the endogenous Pax3 locus and conditional loss of Cdkn2a in Myf6 (myogenic factor 6)-expressing cells. Compared with Pax3PF/PF;Cdkn2aF/F;Myf6ICN/+ controls, Stk3F/F;Stk4F/F;Pax3PF/PF;Cdkn2aF/F;Myf6ICN/+ animals displayed accelerated tumorigenesis (P < 0.0001) and increased tumor penetrance (88% vs. 27%). GEMM tumors were histologically consistent with aRMS. GEMM tumor-derived cell lines showed increased proliferation and invasion and decreased senescence and myogenic differentiation. These data suggest that loss of MST/Hippo signaling acts with Pax3:Foxo1 expression and Cdkn2a loss to promote tumorigenesis. The rapid onset and increased penetrance of tumorigenesis in this model provide a powerful tool for interrogating aRMS biology and screening novel therapeutics.Significance: A novel mouse model sheds light on the critical role of Hippo/MST downregulation in PAX3-FOXO1-positive rhabdomyosarcoma tumorigenesis. Cancer Res; 78(19); 5513-20. ©2018 AACR.

Histotype classification of ovarian carcinoma: A comparison of approaches.

OBJECTIVE: Major changes in the classification of ovarian carcinoma histotypes occurred over the last two decades, resulting in the current 2014 World Health Organization (WHO) diagnostic criteria that recognize five principal histotypes: high-grade serous, low-grade serous, endometrioid, clear cell, and mucinous carcinoma. We assessed the impact of these guidelines and use of immunohistochemical (IHC) markers on classification of ovarian carcinomas in existing population-based studies. METHODS: We evaluated histotype classification for 2361 ovarian carcinomas diagnosed between 1999 and 2009 from two case-control studies using three approaches: 1. pre-2014 WHO ("historic") histotype; 2. Standardized review of pathology slides using the 2014 WHO criteria alone; and 3. An integrated IHC assessment along with the 2014 WHO criteria. We used Kappa statistics to assess agreement between approaches, and Kaplan-Meier survival curves and Cox proportional hazards models to evaluate mortality. RESULTS: Compared to the standardized pathologic review histotype, agreement across approaches was high (kappa = 0.892 for historic, and 0.849 for IHC integrated histotype), but the IHC integrated histotype identified more low-grade serous carcinomas and a subset of endometrioid carcinomas that were assigned as high-grade serous (n = 25). No substantial differences in histotype-specific mortality were observed across approaches. CONCLUSIONS: Our findings suggest that histotype assignment is fairly consistent regardless of classification approach, but that progressive improvements in classification accuracy for some less common histotypes are achieved with pathologic review using the 2014 WHO criteria and with IHC integration. We additionally recommend a classification scheme to fit historic data into the 2014 WHO categories to answer histotype-specific research questions.

Metastatic squamous cell carcinoma with pseudoangiosarcomatous features and aberrant expression of vascular markers.

Squamous cell carcinoma with pseudoangiosarcomatous features is a rare but well-recognized variant of squamous cell carcinoma. These tumors exhibit complex anastomosing channels lined by neoplastic cells, histologically mimicking a vasoformative mesenchymal tumor. Immunohistochemically, the published cases expressed epithelial markers and were consistently negative for vascular markers. Squamous cell carcinoma with pseudoangiosarcomatous features and aberrant expression of vascular markers has never been reported. Herein, we report two cases of metastatic poorly-differentiated squamous cell carcinoma with pseudoangiosarcomatous morphologic features which showed immunoreactivity for vascular markers (CD31, Fli-1, and ERG). One case (left thigh skin squamous cell carcinoma with abdominal wall metastasis) showed strong and diffuse positivity for vascular markers, and the final diagnosis was confirmed with electron microscopy. The second case (squamous cell carcinoma of unknown primary site with bone metastasis) showed patchy positivity for both squamous and vascular markers. This is the first report of squamous cell carcinoma with pseudoangiosarcomatous features and aberrant expression of vascular markers, which resembles angiosarcoma both morphologically and immunohistochemically, and may represent a potential diagnostic pitfall. It is of crucial importance for pathologists to be aware of metastatic squamous cell carcinoma with such unique features, so that misdiagnosis and inappropriate treatment will be avoided.