Profiling endogenous, environmental, and infectious disease mutational signatures in blastic plasmacytoid dendritic cell neoplasms.

BACKGROUND: Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare hematopoietic disease derived from plasmacytoid dendritic lineage cells. The disease typically shows skin as well as frequent bone marrow and peripheral blood involvement. However, the pathogenesis of this disease is still not well understood. While somatic point mutations and genetic rearrangements have been described in BPDCN, the types and origins of these mutations and relationships to other cancer types is not well understood. MATERIALS AND METHODS: To probe the origins of BPDCN, we analyzed the exome sequence data of 9 tumor-normal pair cases of BPDCN. We utilized SignatureAnalyzer, SigProfiler and a custom microbial analysis pipeline to understand the relevance of endogenous and environmental mutagenic processes. RESULTS: Our results identified a significant tobacco exposure and aging genetic signature as well as signatures related to nucleotide excision repair deficiency, ultra violet (UV) exposure, and endogenous deamination in BPDCN. We also assessed the samples for microbial infectious disease organisms but did not find a link to a microbial etiology. CONCLUSION: The identification of a tobacco exposure and aging genetic signature in patients with BPDCN suggests that environmental and endogenous genetic changes may be central to the oncogenesis of BPDCN.

Establishment of a whole slide imaging-based frozen section service at a cancer center.

Background: In recent years, there has been a surge of interest in clinical digital pathology (DP). Hardware and software platforms have matured and become more affordable, and advances in artificial intelligence promise to transform the practice of pathology. At our institution, we are launching a stepwise process of DP adoption which will eventually encompass our entire workflow. Out of necessity, we began by establishing a whole slide imaging (WSI)-based frozen section service. Methods: We proceeded in a systematic manner by first assembling a team of key stakeholders. We carefully evaluated the various options for digitizing frozen sections before deciding that a WSI-based solution made the most sense for us. We used a formalized evaluation system to quantify performance metrics that were relevant to us. After deciding on a WSI-based system, we likewise carefully considered the various whole slide scanners and digital slide management systems available before making decisions. Results: During formal evaluation by pathologists, the WSI-based system outperformed competing platforms. Although implementation was relatively complex, we have been happy with the results and have noticed significant improvements in our frozen section turnaround time. Our users have been happy with the slide management system, which we plan on utilizing in future DP efforts. Conclusions: There are various options for digitizing frozen section slides. Although WSI-based systems are more complex and expensive than some alternatives, they perform well and may make sense for institutions with a pre-existing or planned larger DP infrastructure.

A Study of Thyroid Fine Needle Aspiration of Follicular Adenoma in the "Atypia of Undetermined Significance" Bethesda Category Using Digital Image Analysis.

BACKGROUND: Originally designed for computerized image analysis, ThinPrep is underutilized in that role outside gynecological cytology. It can be used to address the inter/intra-observer variability in the evaluation of thyroid fine needle aspiration (TFNA) biopsy and help pathologists to gain additional insight into thyroid cytomorphology. METHODS: We designed and validated a feature engineering and supervised machine learning-based digital image analysis method using ImageJ and Python scikit-learn . The method was trained and validated from 400 low power (100x) and 400 high power (400x) images generated from 40 TFNA cases. RESULT: The area under the curve (AUC) for receiver operating characteristics (ROC) is 0.75 (0.74-0.82) for model based from low-power images and 0.74 (0.69-0.79) for the model based from high-power images. Cytomorphologic features were synthesized using feature engineering and when performed in isolation, they achieved AUC of 0.71 (0.64-0.77) for chromatin, 0.70 (0.64-0.73) for cellularity, 0.65 (0.60-0.69) for cytoarchitecture, 0.57 (0.51-0.61) for nuclear size, and 0.63 (0.57-0.68) for nuclear shape. CONCLUSION: Our study proves that ThinPrep is an excellent preparation method for digital image analysis of thyroid cytomorphology. It can be used to quantitatively harvest morphologic information for diagnostic purpose.

Autopsy Techniques in Patients With History of Radiofrequency Ablation for Atrial Fibrillation.

CONTEXT.­: Esophageal fistula formation is one of the most feared complications of radiofrequency catheter ablation. This procedure and its many variations, such as the "maze," are becoming the mainstream treatment for atrial fibrillation owing to limitations of antiarrhythmic drugs. The incidence of this complication rate has been reported to be from 0.01% to 1%. OBJECTIVE.­: To delineate the importance of using the en bloc Letulle method of dissection for identifying esophageal fistulas for cases with a history of radiofrequency catheter ablation. DESIGN.­: Six autopsy cases with a history of radiofrequency catheter ablation for atrial fibrillation were selected from 1736 autopsies performed between 2009 and 2020. RESULTS.­: The initial presenting symptoms included neurologic symptoms, chest pains, epigastric discomfort, and sepsis. Transesophageal echocardiogram in 4 cases showed no evidence of thrombus or vegetation, however, 2 cases had evidence of atrial esophageal fistula. The autopsy findings included 5 atrial esophageal fistulas and 1 esophagopericardial fistula. Atrial esophageal fistulas were small and could be detected without difficulty when the en bloc Letulle technique was used and would have been easily missed by the Virchow method. The immediate causes of the deaths were myocardial ischemia, septic emboli to brain and heart, hypovolemic shock secondary to exsanguination, stroke, and coagulopathy. CONCLUSIONS.­: To date, this is the largest collection of autopsy cases showing esophageal fistula associated with prior radiofrequency catheter ablation. The Letulle dissection method is preferable in this setting.

Display Characteristics and Their Impact on Digital Pathology: A Current Review of Pathologists' Future "Microscope".

Digital displays (monitors) are an indispensable component of a pathologists' daily workflow, from writing reports, viewing whole-slide images, or browsing the Internet. Due to a paucity of literature and experience surrounding display use and standardization in pathology, the Food and Drug Administration's (FDA) has currently restricted FDA-cleared whole-slide imaging systems to a specific model of display for each system, which at this time consists of only medical-grade (MG) displays. Further, given that a pathologists' display will essentially become their new surrogate "microscope," it becomes exceedingly important that all pathologists have a basic understanding of fundamental display properties and their functional consequences. This review seeks to: (a) define and summarize the current and emerging display technology, terminology, features, and regulation as they pertain to pathologists and review the current literature on the impact of different display types (e.g. MG vs. consumer off the shelf vs. professional grade) on pathologists' diagnostic performance and (b) discuss the impact of the recent digital pathology device componentization and the coronavirus disease 2019 public emergency on the pixel pathway and display use for remote digital pathology. Display technology has changed dramatically over the past 20 years and continues to change at a rapid rate. There is a paucity of published studies to date that investigate how display type affects pathologist performance, with more research necessary in order to develop standards and minimum specifications for displays in digital pathology. Given the complexity of modern displays, pathologists must become better informed regarding display technology if they wish to have more choice over their future "microscopes."

Artificial Intelligence in Pathology: A Simple and Practical Guide.

Artificial intelligence (AI) is having an increasing impact on the field of pathology, as computation techniques allow computers to perform tasks previously performed by people. Here, we offer a simple and practical guide to AI methods used in pathology, such as digital image analysis, next-generation sequencing, and natural language processing. We not only provide a comprehensive review, but also discuss relevant history and future directions of AI in pathology. We additionally provide a short tabular dictionary of AI terminology which will help practicing pathologists and researchers to understand this field.

Review of different platforms to perform rapid onsite evaluation via telecytology.

There is increased utilisation of cytopathology to provide a rapid onsite evaluation (ROSE) of fine needle aspiration and touch preparations of small biopsies. A well-executed ROSE procedure can significantly impact the diagnostic quality and appropriate specimen triage of procured biopsy materials. To accommodate the demand for ROSE, telecytology has been increasingly implemented to facilitate ROSE occurring remotely. Telecytology can be categorised based on camera systems including eyepiece system, camera port system and robotic microscope/whole slide image scanner system. Image sharing methods include static images, broadcast only live video streaming, teleconferencing and whole slide image management system. In this review, we will discuss the advantages and disadvantages of each of these systems and deployment considerations.

Malignant Melanoma Presenting as Thoracic Midline Malignancy: Clinicopathologic and Molecular Features.

Because of rarity, clinicopathologic and genomic profiles of noncutaneous malignant melanoma (MM) are not well characterized. In this study, we provide a detailed evaluation of 6 cases of MM presenting as thoracic midline malignancy. The tumors occurred in 6 white patients (5 males and 1 female; medium age, 71.5 y; range, 55 to 81 y). At presentation, the disease was confined to the chest (6/6) with 4 cases subsequently showing brain metastasis. All 6 tumors featured epithelioid and 1 case also spindle cell morphology. Three of 6 tumors had melanin and all 6 expressed melanocytic markers. Molecular characterization showed 2 of 6 tumors with BRAF"hot spot" genetic alterations (both with BRAFV600K) and 1 with NF1mutation. All 6 patients died of the disease within 1 year (mean survival, 4.8 mo). One patient showed a partial response to BRAF inhibitor. Our study shows that MM in this location has a spectrum of genetic alterations. The findings highlight the need for comprehensive next generation sequencing-based molecular testing to optimize targeted therapies of this aggressive malignancy.

Comprehensive Study of Telecytology Using Robotic Digital Microscope and Single Z-Stack Digital Scan for Fine-Needle Aspiration-Rapid On-Site Evaluation.

BACKGROUND: The current technology for remote assessment of fine-needle aspiration-rapid on-site evaluation (FNA-ROSE) is limited. Recent advances may provide solutions. This study compared the performance of VisionTek digital microscope (VDM) (Sakura, Japan) and Hamamatsu NanoZoomer C9600-12 single Z-stack digital scan (SZDS) to conventional light microscopy (CLM) for FNA-ROSE. METHODS: We assembled sixty FNA cases from the thyroid (n = 16), lymph node (n = 16), pancreas (n = 9), head and neck (n = 9), salivary gland (n = 5), lung (n = 4), and rectum (n = 1) based on a single institution's routine workflow. One Diff-Quik-stained slide was selected for each case. Two board-certified cytopathologists independently evaluated the cases using VDM, SZDS, and CLM. A "washout" period of at least 14 days was placed between the reviews. The results were categorized into satisfactory versus unsatisfactory for adequacy assessment (AA) and unsatisfactory, benign, atypical, suspicious, and malignant for preliminary diagnosis (PD). RESULTS: For AA, the Cohen's kappa statistics (CKS) scores of intermodality agreement (IMA) were 0.74-0.94 (CLM vs. VDM) and 0.86-1 (CLM vs. SZDS). The discordant rates of IMA were 3.3% (4/120) for VDM versus CLM, and 1.7% (2/120) for SZDS versus CLM. For PD, the CKS scores of IMA ranged 0.7-0.93. The overall discordant rates of IMA were 15% (18/120) for CLM versus VDM and 10.8% (13/120) for CLM versus SZDS. The discordant rates of IMA with 2 or higher degrees were 5.8% (7/120) for CLM versus VDM and 1.7% (2/120) for CLM versus SZDS. The average time spent per slide was 270 s for VDM, significantly longer than that for CLM (113 s) or for SZDS (122 s). CONCLUSIONS: Our data demonstrate that both VDM and SZDS are suitable to provide AA and reasonable PD evaluation. VDM, however, has a significantly longer turnaround time and worse diagnostic performance. The study demonstrates both the potentials and challenges of using VDM and SZDS for FNA-ROSE.

An obesity paradox: an inverse correlation between body mass index and atherosclerosis of the aorta.

BACKGROUND AND AIMS: Morbid obesity generally has been associated with higher morbidity and mortality for a variety of diseases. However, a number of exceptions to this have been reported and referred to as the "obesity paradox." The purpose of the present study was to obtain objective data on aortic atherosclerosis and its relationship to body mass index (BMI, kg/m2), based on autopsy findings in a large cohort of overweight and obese decedents. METHODS: Decedents were ≥18 years who had autopsies between 2003 and 2014, a subset of whom were morbidly obese (BMI≥40). Autopsy findings were reviewed and compared to a control group (BMI<40) who had consecutive autopsies performed between January 2013 and June 2014. Atherosclerosis was assessed by gross pathologic examination using a semiquantitative grading scale (from 0 to 3), and for statistical analysis, the scores were stratified into two groups: nonsevere (<2) or severe (≥2). RESULTS: There were 304 decedents in the study: 66 were morbidly obese (BMI≥40), 94 were either Class I or II obese (BMI 30-40), 127 were either overweight (BMI 25.0-29.9) or normal weight (BMI 20-24.9), and 17 were underweight (BMI<20). Decedents with mild atherosclerosis were significantly younger than those with severe disease (55.2 vs. 67.3, P<.0001). Decedents were further stratified by age and BMI. Univariate analysis revealed that decedents >60 years were more likely to have severe atherosclerosis than those ≤60 years (61% vs. 30%, P<.0001). There was a highly significant (P=.008) inverse relationship between severe aortic atherosclerosis and BMI. Twenty of 66 decedents (30%) with a BMI≥40 had severe atherosclerosis vs. 122 of 238 decedents (51%) with BMIs<40 (P=.001). As BMI increased, the probability of developing severe disease decreased. Hypertension increased the probability of having severe atherosclerosis (54% vs. 33%, P=.007). After adjusting for other covariates, multivariable analysis revealed that age and hypertension were still positively correlated with the severity of atherosclerosis (P=.014 and 0.028, respectively), and the inverse relationship between BMI and atherosclerosis remained (adjusted relative risk of BMI≥40 vs. <40=0.64, 95% confidence interval: 0.4-1; P=.03). CONCLUSIONS: Our data extend the previously described obesity paradox to another disease entity, atherosclerosis of the aorta. Morbid obesity appeared to have a protective effect for developing severe aortic atherosclerosis, for the reasons for which are yet to be determined. However, the mean age at death of decedents with BMIs≥40 was younger than those with BMIs in the 20-30 range (55.9 vs. 63.2 years, P=.001), confirming that morbid obesity was not associated with increased longevity.

Morphometric data on severely and morbidly obese deceased, established on forensic and non-forensic autopsies.

With the widespread increase in the incidence of obesity, autopsies on severely and morbidly obese deceased have become common in the USA. Standard reference tables for organ weights provide little or no information on individuals with a body mass index greater than 35 kg/m(2). Although several recent reports have provided organ weights for small numbers of morbidly obese persons who died naturally from a variety of causes, these data may have been affected by comorbidities. Furthermore, they did not provide information relative to differences in organ weight based on gender, age, and race. The aim of the present study was to fill this void by developing reference tables for organ weights of severely and morbidly obese individuals. Our study was based on data from 802 forensic and medical autopsies, including 435 cases of death of natural and 367 of non-natural causes. Organ weights were compared between these groups, and reference ranges were generated. Significant variability was found in organ weights especially among deceased older than 40 years who died naturally, suggesting that comorbidities affect organ weight. Reference tables were compiled for organ weights and morphometric data based on gender, age, and race. Since obesity is a pathological condition affecting organ weight, these reference tables do not reflect normal organ weights but only weight as seen in severely and morbidly obese individuals. They should be useful to pathologists who perform forensic and non-forensic autopsies.

Profiling critical cancer gene mutations in clinical tumor samples.

BACKGROUND: Detection of critical cancer gene mutations in clinical tumor specimens may predict patient outcomes and inform treatment options; however, high-throughput mutation profiling remains underdeveloped as a diagnostic approach. We report the implementation of a genotyping and validation algorithm that enables robust tumor mutation profiling in the clinical setting. METHODOLOGY: We developed and implemented an optimized mutation profiling platform ("OncoMap") to interrogate approximately 400 mutations in 33 known oncogenes and tumor suppressors, many of which are known to predict response or resistance to targeted therapies. The performance of OncoMap was analyzed using DNA derived from both frozen and FFPE clinical material in a diverse set of cancer types. A subsequent in-depth analysis was conducted on histologically and clinically annotated pediatric gliomas. The sensitivity and specificity of OncoMap were 93.8% and 100% in fresh frozen tissue; and 89.3% and 99.4% in FFPE-derived DNA. We detected known mutations at the expected frequencies in common cancers, as well as novel mutations in adult and pediatric cancers that are likely to predict heightened response or resistance to existing or developmental cancer therapies. OncoMap profiles also support a new molecular stratification of pediatric low-grade gliomas based on BRAF mutations that may have immediate clinical impact. CONCLUSIONS: Our results demonstrate the clinical feasibility of high-throughput mutation profiling to query a large panel of "actionable" cancer gene mutations. In the future, this type of approach may be incorporated into both cancer epidemiologic studies and clinical decision making to specify the use of many targeted anticancer agents.

The 8q24 cancer risk variant rs6983267 shows long-range interaction with MYC in colorectal cancer.

An inherited variant on chromosome 8q24, rs6983267, is significantly associated with cancer pathogenesis. We present evidence that the region harboring this variant is a transcriptional enhancer, that the alleles of rs6983267 differentially bind transcription factor 7-like 2 (TCF7L2) and that the risk region physically interacts with the MYC proto-oncogene. These data provide strong support for a biological mechanism underlying this non-protein-coding risk variant.