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The field of anatomic pathology has been evolving in the last few decades and the advancements have been largely fostered by innovative technology. Immunohistochemistry enabled a paradigm shift in discovery and diagnostic evaluation, followed by booming genomic advancements which allowed for submicroscopic pathologic characterization, and now the field of digital pathology coupled with machine learning and big data acquisition is paving the way to revolutionize the pathology medical domain. Whole slide imaging (WSI) is a disruptive technology where glass slides are digitized to produce on-screen whole slide images. Specifically, in the past decade, there have been significant advances in digital pathology systems that have allowed this technology to promote integration into clinical practice. Whole slide images (WSI), or digital slides, can be viewed and navigated comparable to glass slides on a microscope, as digital files. Whole slide imaging has increased in adoption among pathologists, pathology departments, and scientists for clinical, educational, and research initiatives. Integration of digital pathology systems requires a coordinated effort with numerous stakeholders, not only within the pathology department, but across the entire enterprise. Each pathology department has distinct needs, use cases and blueprints, however the framework components and variables for successful clinical integration can be generalized across any organization seeking to undergo a digital transformation at any scale. This article will review those components and considerations for integrating digital pathology systems into clinical practice.

Thyroid-like follicular renal cell carcinoma is an uncommon kidney tumor with no distinct molecular alteration described to date. This cohort of eight women with mean and median ages of 45 and 46 years, respectively (range 19–65 years), had unencapsulated, well-circumscribed tumors composed of tightly packed anastomosing follicle-like cysts filled with eosinophilic colloid-like material and lined by cuboidal cells with high nuclear to cytoplasmic ratios, oval to elongated nuclei with perpendicular arrangement toward the lumens, and prominent nuclear overlapping. The stroma between these was minimal with the exception of two tumors. Calcifications and necrosis were absent. Immunohistochemically, the tumors were positive for KRT19 (7/7), PAX8 (5/5), cyclin D1 (6/6), KRT7 (5/7), and AMACR (1/5; focal, weak), and were negative for WT1, TTF1 (transcription termination factor-1), and thyroglobulin. In three of three tumors tested molecularly, EWSR1-PATZ1fusion was identified by RNA sequencing and confirmed by RT-PCR and Sanger sequencing. Over a follow-up period of 1–7 years, no evidence of recurrence or metastasis has been detected. The EWSR1-PATZ1fusion has been recognized as a recurrent alteration in a subset of round to spindle cell sarcomas with EWSR1–non-ETSfusions (EWSR1-PATZ1sarcoma) and in several central nervous system tumors. The finding of an EWSR1-PATZ1fusion in all three of the thyroid-like follicular renal cell carcinomas for which sufficient tissue was available for genomic profiling provides the first distinct molecular abnormality in thyroid-like follicular renal cell carcinomas, supporting its designation as a distinct diagnostic entity.

Thyroid-like follicular renal cell carcinoma is an uncommon kidney tumor with no distinct molecular alteration described to date. This cohort of eight women with mean and median ages of 45 and 46 years, respectively (range 19–65 years), had unencapsulated, well-circumscribed tumors composed of tightly packed anastomosing follicle-like cysts filled with eosinophilic colloid-like material and lined by cuboidal cells with high nuclear to cytoplasmic ratios, oval to elongated nuclei with perpendicular arrangement toward the lumens, and prominent nuclear overlapping. The stroma between these was minimal with the exception of two tumors. Calcifications and necrosis were absent. Immunohistochemically, the tumors were positive for KRT19 (7/7), PAX8 (5/5), cyclin D1 (6/6), KRT7 (5/7), and AMACR (1/5; focal, weak), and were negative for WT1, TTF1 (transcription termination factor-1), and thyroglobulin. In three of three tumors tested molecularly, EWSR1-PATZ1fusion was identified by RNA sequencing and confirmed by RT-PCR and Sanger sequencing. Over a follow-up period of 1–7 years, no evidence of recurrence or metastasis has been detected. The EWSR1-PATZ1fusion has been recognized as a recurrent alteration in a subset of round to spindle cell sarcomas with EWSR1–non-ETSfusions (EWSR1-PATZ1sarcoma) and in several central nervous system tumors. The finding of an EWSR1-PATZ1fusion in all three of the thyroid-like follicular renal cell carcinomas for which sufficient tissue was available for genomic profiling provides the first distinct molecular abnormality in thyroid-like follicular renal cell carcinomas, supporting its designation as a distinct diagnostic entity.

The field of anatomic pathology has been evolving in the last few decades and the advancements have been largely fostered by innovative technology. Immunohistochemistry enabled a paradigm shift in discovery and diagnostic evaluation, followed by booming genomic advancements which allowed for submicroscopic pathologic characterization, and now the field of digital pathology coupled with machine learning and big data acquisition is paving the way to revolutionize the pathology medical domain. Whole slide imaging (WSI) is a disruptive technology where glass slides are digitized to produce on-screen whole slide images. Specifically, in the past decade, there have been significant advances in digital pathology systems that have allowed this technology to promote integration into clinical practice. Whole slide images (WSI), or digital slides, can be viewed and navigated comparable to glass slides on a microscope, as digital files. Whole slide imaging has increased in adoption among pathologists, pathology departments, and scientists for clinical, educational, and research initiatives. Integration of digital pathology systems requires a coordinated effort with numerous stakeholders, not only within the pathology department, but across the entire enterprise. Each pathology department has distinct needs, use cases and blueprints, however the framework components and variables for successful clinical integration can be generalized across any organization seeking to undergo a digital transformation at any scale. This article will review those components and considerations for integrating digital pathology systems into clinical practice.

In the modern era, detailed pathologic characteristics of a thyroid tumor are crucial to achieve accurate diagnosis and guide treatment. The presence of capsular invasion (CI) is diagnostic for carcinoma, whereas vascular invasion (VI) and nodal metastasis (NM) are included in risk stratification. However, the very definition of CI and VI is surrounded by controversies and an accurate assessment of NM is lacking. Whole Block Imaging (WBI) by microCT is a new imaging modality to create 3D reconstruction of whole tissue block with microscopic level resolution without the need for tissue sectioning. In this study, we aimed to define CI, VI, and NM volume using WBI by microCT. Twenty-eight paraffin blocks (PBs) from 26 thyroid tumors were scanned. Ten PBs contained CI, whereas 7 had VI. 3D microCT images were compared with whole slide images (WSI) of corresponding H&E slides. In 2 cases with VI and/or CI, WSI of serial H&E slides were obtained and underwent 3D-reconstruction to be compared with the WBI. Satellite tumor nodules beyond tumor capsule were shown to be CI by demonstrating the point of penetration using microCT and 3D reconstruction. Additional foci of CI were detected using microCT. VI was seen using microCT. Fibrin associated with tumor thrombus was not always present on serially sectioned H&E slides. WBI by microCT scanner was able to assess the volume of NM. In conclusion, WBI is able to detect CI, VI, and assess the volume of NM in thyroid carcinoma without tissue sectioning. It is the ultimate method for the complete sampling of the tumor capsule. It has the potential to increase the detection rate of CI, better define criteria for CI and VI, and provide an accurate assessment of the volume of nodal disease. This technology may impact the future practice of surgical pathology.

In the modern era, detailed pathologic characteristics of a thyroid tumor are crucial to achieve accurate diagnosis and guide treatment. The presence of capsular invasion (CI) is diagnostic for carcinoma, whereas vascular invasion (VI) and nodal metastasis (NM) are included in risk stratification. However, the very definition of CI and VI is surrounded by controversies and an accurate assessment of NM is lacking. Whole Block Imaging (WBI) by microCT is a new imaging modality to create 3D reconstruction of whole tissue block with microscopic level resolution without the need for tissue sectioning. In this study, we aimed to define CI, VI, and NM volume using WBI by microCT. Twenty-eight paraffin blocks (PBs) from 26 thyroid tumors were scanned. Ten PBs contained CI, whereas 7 had VI. 3D microCT images were compared with whole slide images (WSI) of corresponding H&E slides. In 2 cases with VI and/or CI, WSI of serial H&E slides were obtained and underwent 3D-reconstruction to be compared with the WBI. Satellite tumor nodules beyond tumor capsule were shown to be CI by demonstrating the point of penetration using microCT and 3D reconstruction. Additional foci of CI were detected using microCT. VI was seen using microCT. Fibrin associated with tumor thrombus was not always present on serially sectioned H&E slides. WBI by microCT scanner was able to assess the volume of NM. In conclusion, WBI is able to detect CI, VI, and assess the volume of NM in thyroid carcinoma without tissue sectioning. It is the ultimate method for the complete sampling of the tumor capsule. It has the potential to increase the detection rate of CI, better define criteria for CI and VI, and provide an accurate assessment of the volume of nodal disease. This technology may impact the future practice of surgical pathology.

Remote digital pathology allows healthcare systems to maintain pathology operations during public health emergencies. Existing Clinical Laboratory Improvement Amendments regulations require pathologists to electronically verify patient reports from a certified facility. During the 2019 pandemic of COVID-19 disease, caused by the SAR-CoV-2 virus, this requirement potentially exposes pathologists, their colleagues, and household members to the risk of becoming infected. Relaxation of government enforcement of this regulation allows pathologists to review and report pathology specimens from a remote, non-CLIA certified facility. The availability of digital pathology systems can facilitate remote microscopic diagnosis, although formal comprehensive (case-based) validation of remote digital diagnosis has not been reported. All glass slides representing routine clinical signout workload in surgical pathology subspecialties at Memorial Sloan Kettering Cancer Center were scanned on an Aperio GT450 at ×40 equivalent resolution (0.26 µm/pixel). Twelve pathologists from nine surgical pathology subspecialties remotely reviewed and reported complete pathology cases using a digital pathology system from a non-CLIA certified facility through a secure connection. Whole slide images were integrated to and launched within the laboratory information system to a custom vendor-agnostic, whole slide image viewer. Remote signouts utilized consumer-grade computers and monitors (monitor size, 13.3–42 in.; resolution, 1280 × 800–3840 × 2160 pixels) connecting to an institution clinical workstation via secure virtual private network. Pathologists subsequently reviewed all corresponding glass slides using a light microscope within the CLIA-certified department. Intraobserver concordance metrics included reporting elements of top-line diagnosis, margin status, lymphovascular and/or perineural invasion, pathology stage, and ancillary testing. The median whole slide image file size was 1.3 GB; scan time/slide averaged 90 s; and scanned tissue area averaged 612 mm2. Signout sessions included a total of 108 cases, comprised of 254 individual parts and 1196 slides. Major diagnostic equivalency was 100% between digital and glass slide diagnoses; and overall concordance was 98.8% (251/254). This study reports validation of primary diagnostic review and reporting of complete pathology cases from a remote site during a public health emergency. Our experience shows high (100%) intraobserver digital to glass slide major diagnostic concordance when reporting from a remote site. This randomized, prospective study successfully validated remote use of a digital pathology system including operational feasibility supporting remote review and reporting of pathology specimens, and evaluation of remote access performance and usability for remote signout.

Remote digital pathology allows healthcare systems to maintain pathology operations during public health emergencies. Existing Clinical Laboratory Improvement Amendments regulations require pathologists to electronically verify patient reports from a certified facility. During the 2019 pandemic of COVID-19 disease, caused by the SAR-CoV-2 virus, this requirement potentially exposes pathologists, their colleagues, and household members to the risk of becoming infected. Relaxation of government enforcement of this regulation allows pathologists to review and report pathology specimens from a remote, non-CLIA certified facility. The availability of digital pathology systems can facilitate remote microscopic diagnosis, although formal comprehensive (case-based) validation of remote digital diagnosis has not been reported. All glass slides representing routine clinical signout workload in surgical pathology subspecialties at Memorial Sloan Kettering Cancer Center were scanned on an Aperio GT450 at ×40 equivalent resolution (0.26 µm/pixel). Twelve pathologists from nine surgical pathology subspecialties remotely reviewed and reported complete pathology cases using a digital pathology system from a non-CLIA certified facility through a secure connection. Whole slide images were integrated to and launched within the laboratory information system to a custom vendor-agnostic, whole slide image viewer. Remote signouts utilized consumer-grade computers and monitors (monitor size, 13.3–42 in.; resolution, 1280 × 800–3840 × 2160 pixels) connecting to an institution clinical workstation via secure virtual private network. Pathologists subsequently reviewed all corresponding glass slides using a light microscope within the CLIA-certified department. Intraobserver concordance metrics included reporting elements of top-line diagnosis, margin status, lymphovascular and/or perineural invasion, pathology stage, and ancillary testing. The median whole slide image file size was 1.3 GB; scan time/slide averaged 90 s; and scanned tissue area averaged 612 mm2. Signout sessions included a total of 108 cases, comprised of 254 individual parts and 1196 slides. Major diagnostic equivalency was 100% between digital and glass slide diagnoses; and overall concordance was 98.8% (251/254). This study reports validation of primary diagnostic review and reporting of complete pathology cases from a remote site during a public health emergency. Our experience shows high (100%) intraobserver digital to glass slide major diagnostic concordance when reporting from a remote site. This randomized, prospective study successfully validated remote use of a digital pathology system including operational feasibility supporting remote review and reporting of pathology specimens, and evaluation of remote access performance and usability for remote signout.

Herein we described the clinical, radiological, histological, and molecular characteristics of seven soft tissue aneurysmal bone cysts (STABCs) diagnosed and managed at a tertiary cancer center and to elucidate their relationship with myositis ossificans (MO). All cases had established imaging and histopathological diagnosis of STABC and were subject to fluorescence in situ hybridization (FISH) for USP6rearrangement and Archer® FusionPlex® targeted RNA sequencing (RNASeq) analysis to identify the fusion partner. A thorough literature review of STABC and MO was conducted. The patients presented with painful masses unpreceded by trauma, occurring most commonly in the deep soft tissue of the thigh/gluteus (4/7), and also in the supraclavicular region, the axilla, and the hand. On imaging, the lesions were frequently associated with peripheral calcification on conventional radiographs and CT (6/7), cystic components on ultrasound, as well as perilesional edema (7/7) and fluid levels (3/7) on MRI. Bone scan (1/1) showed intense radiotracer uptake. Histologically, 6/7 cases demonstrated zonal arrangements reminiscent of MO. USP6rearrangement was found in all seven cases by FISH and/or RNASeq. RNASeq further detected COL1A1-USP6fusion in six cases and a novel ANGPTL2-USP6fusion in one case. Four patients underwent resection of the tumors and were disease free at their last follow-up. Three patients who underwent incisional or needle biopsies had no evidence of disease progression on imaging studies. In conclusion, the clinical, radiological, and pathological overlap between STABC and MO suggests that they are closely related entities. A novel fusion ANGPTL2-USP6is associated with distinct clinical and pathological presentation.

Herein we described the clinical, radiological, histological, and molecular characteristics of seven soft tissue aneurysmal bone cysts (STABCs) diagnosed and managed at a tertiary cancer center and to elucidate their relationship with myositis ossificans (MO). All cases had established imaging and histopathological diagnosis of STABC and were subject to fluorescence in situ hybridization (FISH) for USP6rearrangement and Archer® FusionPlex® targeted RNA sequencing (RNASeq) analysis to identify the fusion partner. A thorough literature review of STABC and MO was conducted. The patients presented with painful masses unpreceded by trauma, occurring most commonly in the deep soft tissue of the thigh/gluteus (4/7), and also in the supraclavicular region, the axilla, and the hand. On imaging, the lesions were frequently associated with peripheral calcification on conventional radiographs and CT (6/7), cystic components on ultrasound, as well as perilesional edema (7/7) and fluid levels (3/7) on MRI. Bone scan (1/1) showed intense radiotracer uptake. Histologically, 6/7 cases demonstrated zonal arrangements reminiscent of MO. USP6rearrangement was found in all seven cases by FISH and/or RNASeq. RNASeq further detected COL1A1-USP6fusion in six cases and a novel ANGPTL2-USP6fusion in one case. Four patients underwent resection of the tumors and were disease free at their last follow-up. Three patients who underwent incisional or needle biopsies had no evidence of disease progression on imaging studies. In conclusion, the clinical, radiological, and pathological overlap between STABC and MO suggests that they are closely related entities. A novel fusion ANGPTL2-USP6is associated with distinct clinical and pathological presentation.

GLI1(12q13.3) amplification is identified in a subset of mesenchymal neoplasms with a distinct nested round cell/epithelioid phenotype. MDM2and CDK4genes are situated along the oncogenic 12q13-15 segment, amplification of which defines well-differentiated (WDLPS)/dedifferentiated liposarcoma (DDLPS). The 12q amplicon can occasionally include GLI1– a gene in close proximity to CDK4. We hereby describe the first cohort of GLI1/MDM2/CDK4co-amplified WD/DDLPS.

Whole slide imaging is Food and Drug Administration-approved for primary diagnosis in the United States of America; however, relatively few pathology departments in the country have fully implemented an enterprise wide digital pathology system enabled for primary diagnosis. Digital pathology has significant potential to transform pathology practice with several published studies documenting some level of diagnostic equivalence between digital and conventional systems. However, whole slide imaging also has significant potential to disrupt pathology practice, due to the differences in efficiency of manipulating digital images vis-à-vis glass slides, and studies on the efficiency of actual digital pathology workload are lacking. Our randomized, equivalency and efficiency study aimed to replicate clinical workflow, comparing conventional microscopy to a complete digital pathology signout using whole slide images, evaluating the equivalency and efficiency of glass slide to whole slide image reporting, reflective of true pathology practice workloads in the clinical setting. All glass slides representing an entire day's routine clinical signout workload for six different anatomic pathology subspecialties at Memorial Sloan Kettering Cancer Center were scanned on Leica Aperio AT2 at ×40 (0.25 µm/pixel). Integration of whole slide images for each accessioned case is through an interface between the Leica eSlide manager database and the laboratory information system, Cerner CoPathPlus. Pathologists utilized a standard institution computer workstation and viewed whole slide images through an internally developed, vendor agnostic whole slide image viewer, named the “MSK Slide Viewer”. Subspecialized pathologists first reported on glass slides from surgical pathology cases using routine clinical workflow. Glass slides were de-identified, scanned, and re-accessioned in the laboratory information system test environment. After a washout period of 13 weeks, pathologists reported the same clinical workload using whole slide image integrated within the laboratory information system. Intraobserver equivalency metrics included top-line diagnosis, margin status, lymphovascular and/or perineural invasion, pathology stage, and the need to order ancillary testing (i.e., recuts, immunohistochemistry). Turnaround time (efficiency) evaluation was defined by the start of each case when opened in the laboratory information system and when the case was completed for that day (i.e., case sent to signout queue or pending ancillary studies). Eight pathologists participated from the following subspecialties: bone and soft tissue, genitourinary, gastrointestinal, breast, gynecologic, and dermatopathology. Glass slides signouts comprised of 204 cases, encompassing 2091 glass slides; and digital signouts comprised of 199 cases, encompassing 2073 whole slide images. The median whole slide image file size was 1.54 GB; scan time/slide, 6 min 24 s; and scan area 32.1 × 18.52 mm. Overall diagnostic equivalency (e.g., top-line diagnosis) was 99.3% between digital and glass slide signout; however, signout using whole slide images showed a median overall 19% decrease in efficiency per case. No significant difference by reader, subspecialty, or specimen type was identified. Our experience is the most comprehensive study to date and shows high intraobserver whole slide image to glass slide equivalence in reporting of true clinical workflows and workloads. Efficiency needs to improve for digital pathology to gain more traction among pathologists.

Whole slide imaging is Food and Drug Administration-approved for primary diagnosis in the United States of America; however, relatively few pathology departments in the country have fully implemented an enterprise wide digital pathology system enabled for primary diagnosis. Digital pathology has significant potential to transform pathology practice with several published studies documenting some level of diagnostic equivalence between digital and conventional systems. However, whole slide imaging also has significant potential to disrupt pathology practice, due to the differences in efficiency of manipulating digital images vis-à-vis glass slides, and studies on the efficiency of actual digital pathology workload are lacking. Our randomized, equivalency and efficiency study aimed to replicate clinical workflow, comparing conventional microscopy to a complete digital pathology signout using whole slide images, evaluating the equivalency and efficiency of glass slide to whole slide image reporting, reflective of true pathology practice workloads in the clinical setting. All glass slides representing an entire day’s routine clinical signout workload for six different anatomic pathology subspecialties at Memorial Sloan Kettering Cancer Center were scanned on Leica Aperio AT2 at ×40 (0.25 µm/pixel). Integration of whole slide images for each accessioned case is through an interface between the Leica eSlide manager database and the laboratory information system, Cerner CoPathPlus. Pathologists utilized a standard institution computer workstation and viewed whole slide images through an internally developed, vendor agnostic whole slide image viewer, named the “MSK Slide Viewer”. Subspecialized pathologists first reported on glass slides from surgical pathology cases using routine clinical workflow. Glass slides were de-identified, scanned, and re-accessioned in the laboratory information system test environment. After a washout period of 13 weeks, pathologists reported the same clinical workload using whole slide image integrated within the laboratory information system. Intraobserver equivalency metrics included top-line diagnosis, margin status, lymphovascular and/or perineural invasion, pathology stage, and the need to order ancillary testing (i.e., recuts, immunohistochemistry). Turnaround time (efficiency) evaluation was defined by the start of each case when opened in the laboratory information system and when the case was completed for that day (i.e., case sent to signout queue or pending ancillary studies). Eight pathologists participated from the following subspecialties: bone and soft tissue, genitourinary, gastrointestinal, breast, gynecologic, and dermatopathology. Glass slides signouts comprised of 204 cases, encompassing 2091 glass slides; and digital signouts comprised of 199 cases, encompassing 2073 whole slide images. The median whole slide image file size was 1.54 GB; scan time/slide, 6 min 24 s; and scan area 32.1 × 18.52 mm. Overall diagnostic equivalency (e.g., top-line diagnosis) was 99.3% between digital and glass slide signout; however, signout using whole slide images showed a median overall 19% decrease in efficiency per case. No significant difference by reader, subspecialty, or specimen type was identified. Our experience is the most comprehensive study to date and shows high intraobserver whole slide image to glass slide equivalence in reporting of true clinical workflows and workloads. Efficiency needs to improve for digital pathology to gain more traction among pathologists.

Integration of morphological, immunohistochemical, and molecular methods is often necessary for the precise diagnosis and optimal clinical management of sarcomas. We have validated and implemented a clinical molecular diagnostic assay, MSK- Fusion Solid, for detection of gene fusions in solid tumors, including sarcomas. Starting with RNA extracted from formalin-fixed paraffin-embedded tumor material, this targeted RNA sequencing assay utilizes anchored multiplex PCR to detect oncogenic fusion transcripts involving 62 genes known to be recurrently rearranged in solid tumors including sarcomas without prior knowledge of fusion partners. From 1/2016 to 1/2018, 192 bone and soft tissue tumors were submitted for MSK- Fusion Solid analysis and 96% (184/192) successfully passed all the pre-sequencing quality control parameters and sequencing steps. These sarcomas encompass 24 major tumor types, including 175 soft tissue tumors and 9 osteosarcomas. Ewing and Ewing-like sarcomas, rhabdomyosarcoma, and sarcoma-not otherwise specified were the three most common tumor types. Diagnostic in-frame fusion transcripts were detected in 43% of cases, including 3% (6/184) with novel fusion partners, specifically TRPS1-PLAG1, VCP-TFE3, MYLK-BRAF, FUS-TFCP2, and ACTB-FOSB, the latter in two cases of pseudomyogenic hemangioendothelioma, representing a novel observation in this sarcoma. Our experience shows that this targeted RNA sequencing assay performs in a robust and sensitive fashion on RNA extracted from most routine clinical specimens of sarcomas thereby facilitating precise diagnosis and providing opportunities for novel fusion partner discovery.

Integration of morphological, immunohistochemical, and molecular methods is often necessary for the precise diagnosis and optimal clinical management of sarcomas. We have validated and implemented a clinical molecular diagnostic assay, MSK- Fusion Solid, for detection of gene fusions in solid tumors, including sarcomas. Starting with RNA extracted from formalin-fixed paraffin-embedded tumor material, this targeted RNA sequencing assay utilizes anchored multiplex PCR to detect oncogenic fusion transcripts involving 62 genes known to be recurrently rearranged in solid tumors including sarcomas without prior knowledge of fusion partners. From 1/2016 to 1/2018, 192 bone and soft tissue tumors were submitted for MSK- Fusion Solid analysis and 96% (184/192) successfully passed all the pre-sequencing quality control parameters and sequencing steps. These sarcomas encompass 24 major tumor types, including 175 soft tissue tumors and 9 osteosarcomas. Ewing and Ewing-like sarcomas, rhabdomyosarcoma, and sarcoma-not otherwise specified were the three most common tumor types. Diagnostic in-frame fusion transcripts were detected in 43% of cases, including 3% (6/184) with novel fusion partners, specifically TRPS1-PLAG1, VCP-TFE3, MYLK-BRAF, FUS-TFCP2, and ACTB-FOSB, the latter in two cases of pseudomyogenic hemangioendothelioma, representing a novel observation in this sarcoma. Our experience shows that this targeted RNA sequencing assay performs in a robust and sensitive fashion on RNA extracted from most routine clinical specimens of sarcomas thereby facilitating precise diagnosis and providing opportunities for novel fusion partner discovery.

IDH2R172 mutations occur in >80% sinonasal undifferentiated carcinomas (“SNUC”) and ~80% of these are R172S and R172T variants. We examined the utility of the monoclonal antibody 11C8B1 to IDH2R172S in IDH2R172-mutated tumors to establish an immunohistochemistry protocol as a surrogate method for IDH2R172S mutation detection. Eighty-eight formalin-fixed paraffin-embedded tumors including 42 sinonasal tumors and a variety of IDH1/2-mutated malignancies were tested by immunohistochemistry. The IDH1/2mutation status was determined in 86 cases by a targeted massively parallel sequencing MSK-IMPACTTMassay. Interestingly, monoclonal antibody 11C8B1 was reactive with all IDH2R172S (N= 15) mutated tumors including 12 sinonasal carcinomas, 2 high-grade sarcomas and one intrahepatic cholangiocarcinoma, and with all R172T (N= 3) mutated sinonasal carcinomas displaying a distinct granular cytoplasmic labeling in all R172S/T mutated malignancies. 11C8B1 immunohistochemistry was also positive in 2 of 6 IDH1R132S-mutated tumors, including one intrahepatic cholangiocarcinoma and one chondrosarcoma showing a smooth homogeneous cytoplasmic staining pattern. All IDH2R172G/K/M/W (N= 22) and IDH1132H/C/G/L (N= 15) mutated tumors, and all IDH1/2-wild-type tumors (N= 25), including a histologic variety of 23 sinonasal tumors, were immunonegative. Importantly, 11 sinonasal undifferentiated carcinomas (N= 14, 79%) and 3 (100%) high-grade neuroendocrine carcinomas, large cell type were 11C8B1 immunopositive. Literature search revealed a virtual absence of IDH2R172 and IDH1R132S mutations in >1000 cases of 8 different malignancies included in the differential diagnosis of sinonasal undifferentiated carcinoma. Our study suggests that positive IDH2 11C8B1 immunohistochemistry in sinonasal carcinomas would be highly predictive of the presence of IDH2R172S/T mutations and could serve as a reliable adjunct diagnostic marker of sinonasal undifferentiated carcinomas in >70% cases.

IDH2R172 mutations occur in >80% sinonasal undifferentiated carcinomas (“SNUC”) and ~80% of these are R172S and R172T variants. We examined the utility of the monoclonal antibody 11C8B1 to IDH2R172S in IDH2R172-mutated tumors to establish an immunohistochemistry protocol as a surrogate method for IDH2R172S mutation detection. Eighty-eight formalin-fixed paraffin-embedded tumors including 42 sinonasal tumors and a variety of IDH1/2-mutated malignancies were tested by immunohistochemistry. The IDH1/2mutation status was determined in 86 cases by a targeted massively parallel sequencing MSK-IMPACTTMassay. Interestingly, monoclonal antibody 11C8B1 was reactive with all IDH2R172S (N= 15) mutated tumors including 12 sinonasal carcinomas, 2 high-grade sarcomas and one intrahepatic cholangiocarcinoma, and with all R172T (N= 3) mutated sinonasal carcinomas displaying a distinct granular cytoplasmic labeling in all R172S/T mutated malignancies. 11C8B1 immunohistochemistry was also positive in 2 of 6 IDH1R132S-mutated tumors, including one intrahepatic cholangiocarcinoma and one chondrosarcoma showing a smooth homogeneous cytoplasmic staining pattern. All IDH2R172G/K/M/W (N= 22) and IDH1132H/C/G/L (N= 15) mutated tumors, and all IDH1/2-wild-type tumors (N= 25), including a histologic variety of 23 sinonasal tumors, were immunonegative. Importantly, 11 sinonasal undifferentiated carcinomas (N= 14, 79%) and 3 (100%) high-grade neuroendocrine carcinomas, large cell type were 11C8B1 immunopositive. Literature search revealed a virtual absence of IDH2R172 and IDH1R132S mutations in >1000 cases of 8 different malignancies included in the differential diagnosis of sinonasal undifferentiated carcinoma. Our study suggests that positive IDH2 11C8B1 immunohistochemistry in sinonasal carcinomas would be highly predictive of the presence of IDH2R172S/T mutations and could serve as a reliable adjunct diagnostic marker of sinonasal undifferentiated carcinomas in >70% cases.

Erdheim–Chester disease is a rare, non-Langerhans cell histiocytosis histologically characterized by multi-systemic proliferation of mature histiocytes in a background of inflammatory stroma. The disease can involve virtually any organ system; most commonly the bones, skin, retroperitoneum, heart, orbit, lung, and brain are affected. Although a histiocytic proliferation is the histological hallmark of the disease, a wide range of morphological appearances have been described as part of case studies or small series. A comprehensive review of histopathological features in clinically and molecularly defined Erdheim–Chester disease has yet to be characterized. To address this issue and help guide clinical practice, we comprehensively analyzed the pathological spectrum of Erdheim–Chester disease in a clinically and molecularly defined cohort. We reviewed 73 biopsies from 42 patients showing involvement by histiocytosis from a variety of organ systems, including bone (16), retroperitoneum (11), skin (19), orbit (6), brain (5), lung (6), cardiac structures (2), epidural soft tissue (3), oral cavity (2), subcutaneous soft tissue (2), and testis (2). In eight patients, one or more bone marrow biopsies were performed due to clinical indication and an accompanying myeloid neoplasm was detected in six of them. Thirty-eight cases were investigated for genetic abnormalities. Somatic mutations involving BRAF(25/38), MAP2K1(6/38), ARAF(2/38), MAP2K2(1/38), KRAS(1/38), and NRAS(1/38) genes were detected. One of the cases with a MAP2K1mutation also harbored a PIK3CAmutation. We have observed marked heterogeneity in histology and immunophenotype, identified site-specific features, overlap with other histiocytic and myeloid disorders and potential diagnostic pitfalls. We hope that broadening the spectrum of recognized pathologic manifestations of Erdheim–Chester disease will help practicing clinicians and pathologists to diagnose Erdheim–Chester disease early in the disease course and manage these patients effectively.

High-grade endometrial stromal sarcoma likely encompasses underrecognized tumors harboring genetic abnormalities besides YWHAE–NUTM2fusion. Triggered by three initial endometrial stromal sarcomas with ZC3H7B–BCORfusion characterized by high-grade morphology and aggressive clinical behavior, we herein investigate the clinicopathologic features of this genetic subset by expanding the analysis to 17 such tumors. All of them occurred in adult women with a median age of 54 (range, 28–71) years. They were predominantly based in the endomyometrium and demonstrated tongue-like and/or pushing myometrial invasion. Most were uniformly cellular and displayed haphazard fascicles of spindle cells with mild to moderate nuclear atypia. Myxoid matrix was seen in 14 of 17 (82%) tumors, and collagen plaques were seen in 8 (47%). The mitotic index was ≥10 mitotic figures/10 high-power fields (HPFs) in 14 of 17 (82%) tumors with a median of 14.5 mitotic figures/10 HPFs. No foci of conventional or variant low-grade endometrial stromal sarcoma were seen. All tumors expressed CD10 with only limited or absent desmin, SMA and/or h-caldesmon staining. ER and PR expression in >5% of cells was seen in 4 of 12 (33%) tumors. Diffuse cyclin D1 and BCOR immunoreactivity was present in 7 of 8 (88%) and 7 of 14 (50%) tumors, respectively. Fluorescence in situhybridization or targeted RNA sequencing confirmed ZC3H7B–BCORfusion in all tumors, including four and two previously diagnosed as myxoid leiomyosarcoma and undifferentiated uterine sarcoma, respectively. Limited clinical data suggest that patients present at higher stage and have worse prognosis compared with published outcomes in low-grade endometrial stromal sarcoma. Tumors with ZC3H7B–BCORfusion constitute a distinct group of endometrial stromal sarcomas with high-grade morphology that should be distinguished from other uterine mesenchymal neoplasms that may demonstrate myxoid morphology.

High-grade endometrial stromal sarcoma likely encompasses underrecognized tumors harboring genetic abnormalities besides YWHAE–NUTM2fusion. Triggered by three initial endometrial stromal sarcomas with ZC3H7B–BCORfusion characterized by high-grade morphology and aggressive clinical behavior, we herein investigate the clinicopathologic features of this genetic subset by expanding the analysis to 17 such tumors. All of them occurred in adult women with a median age of 54 (range, 28–71) years. They were predominantly based in the endomyometrium and demonstrated tongue-like and/or pushing myometrial invasion. Most were uniformly cellular and displayed haphazard fascicles of spindle cells with mild to moderate nuclear atypia. Myxoid matrix was seen in 14 of 17 (82%) tumors, and collagen plaques were seen in 8 (47%). The mitotic index was ≥10 mitotic figures/10 high-power fields (HPFs) in 14 of 17 (82%) tumors with a median of 14.5 mitotic figures/10 HPFs. No foci of conventional or variant low-grade endometrial stromal sarcoma were seen. All tumors expressed CD10 with only limited or absent desmin, SMA and/or h-caldesmon staining. ER and PR expression in >5% of cells was seen in 4 of 12 (33%) tumors. Diffuse cyclin D1 and BCOR immunoreactivity was present in 7 of 8 (88%) and 7 of 14 (50%) tumors, respectively. Fluorescence in situhybridization or targeted RNA sequencing confirmed ZC3H7B–BCORfusion in all tumors, including four and two previously diagnosed as myxoid leiomyosarcoma and undifferentiated uterine sarcoma, respectively. Limited clinical data suggest that patients present at higher stage and have worse prognosis compared with published outcomes in low-grade endometrial stromal sarcoma. Tumors with ZC3H7B–BCORfusion constitute a distinct group of endometrial stromal sarcomas with high-grade morphology that should be distinguished from other uterine mesenchymal neoplasms that may demonstrate myxoid morphology.

Erdheim–Chester disease is a rare, non-Langerhans cell histiocytosis histologically characterized by multi-systemic proliferation of mature histiocytes in a background of inflammatory stroma. The disease can involve virtually any organ system; most commonly the bones, skin, retroperitoneum, heart, orbit, lung, and brain are affected. Although a histiocytic proliferation is the histological hallmark of the disease, a wide range of morphological appearances have been described as part of case studies or small series. A comprehensive review of histopathological features in clinically and molecularly defined Erdheim–Chester disease has yet to be characterized. To address this issue and help guide clinical practice, we comprehensively analyzed the pathological spectrum of Erdheim–Chester disease in a clinically and molecularly defined cohort. We reviewed 73 biopsies from 42 patients showing involvement by histiocytosis from a variety of organ systems, including bone (16), retroperitoneum (11), skin (19), orbit (6), brain (5), lung (6), cardiac structures (2), epidural soft tissue (3), oral cavity (2), subcutaneous soft tissue (2), and testis (2). In eight patients, one or more bone marrow biopsies were performed due to clinical indication and an accompanying myeloid neoplasm was detected in six of them. Thirty-eight cases were investigated for genetic abnormalities. Somatic mutations involving BRAF(25/38), MAP2K1(6/38), ARAF(2/38), MAP2K2(1/38), KRAS(1/38), and NRAS(1/38) genes were detected. One of the cases with a MAP2K1mutation also harbored a PIK3CAmutation. We have observed marked heterogeneity in histology and immunophenotype, identified site-specific features, overlap with other histiocytic and myeloid disorders and potential diagnostic pitfalls. We hope that broadening the spectrum of recognized pathologic manifestations of Erdheim–Chester disease will help practicing clinicians and pathologists to diagnose Erdheim–Chester disease early in the disease course and manage these patients effectively.

Recognition of high-grade endometrial stromal sarcoma is important because of its aggressive clinical behavior. Morphologic features of YWHAE-NUTM2 high-grade endometrial stromal sarcoma may overlap with other uterine sarcoma types. BCOR immunoexpression was studied in these tumors and their morphologic mimics to assess its diagnostic utility. BCOR immunohistochemical staining was performed on archival tissue from 28 high-grade endometrial stromal sarcomas with classic morphology (20 YWHAE-NUTM2, 5 ZC3H7B-BCOR, 3 BCOR-ZC3H7B), 3 high-grade endometrial stromal sarcomas with unusual morphology and unknown gene rearrangement status, 66 low-grade endometrial stromal sarcomas, 21 endometrial stromal nodules, 38 uterine leiomyosarcomas, and 19 uterine leiomyomas. Intensity of nuclear staining and percentage of positive tumor cells were recorded. Strong diffuse nuclear BCOR staining (defined as >95% of tumor cells) was seen in the round cell component of all 20 (100%) classic YWHAE-NUTM2 high-grade endometrial stromal sarcomas and the 3 unusual high-grade endometrial stromal sarcomas which prompted FISH studies confirming YWHAE rearrangement in 2 tumors. Genomic PCR confirmed the presence of BCOR exon 16 internal tandem duplication in the third case. Diffuse BCOR staining was strong in three and weak in one BCOR-rearranged high-grade endometrial stromal sarcoma while absent in the remaining four BCOR-rearranged tumors. BCOR staining was weakly positive in <5% of tumor cells in 4 of 66 (6%) low-grade endometrial stromal sarcomas and 1 of 18 (6%) endometrial stromal nodules and weakly to moderately positive in <5–40% of tumor cells in 6 of 31 (19%) leiomyosarcomas. No BCOR staining was seen in the remaining low-grade endometrial stromal sarcomas, endometrial stromal nodules, leiomyosarcomas, or any of the leiomyomas. BCOR immunohistochemical staining is a highly sensitive marker for YWHAE-NUTM2 high-grade endometrial stromal sarcoma with both classic and unusual morphology and identifies a subset of high-grade endometrial stromal sarcoma with BCOR alterations, including BCOR rearrangement and internal tandem duplication.

Recognition of high-grade endometrial stromal sarcoma is important because of its aggressive clinical behavior. Morphologic features of YWHAE-NUTM2high-grade endometrial stromal sarcoma may overlap with other uterine sarcoma types. BCOR immunoexpression was studied in these tumors and their morphologic mimics to assess its diagnostic utility. BCOR immunohistochemical staining was performed on archival tissue from 28 high-grade endometrial stromal sarcomas with classic morphology (20 YWHAE-NUTM2, 5 ZC3H7B-BCOR, 3 BCOR-ZC3H7B), 3 high-grade endometrial stromal sarcomas with unusual morphology and unknown gene rearrangement status, 66 low-grade endometrial stromal sarcomas, 21 endometrial stromal nodules, 38 uterine leiomyosarcomas, and 19 uterine leiomyomas. Intensity of nuclear staining and percentage of positive tumor cells were recorded. Strong diffuse nuclear BCOR staining (defined as >95% of tumor cells) was seen in the round cell component of all 20 (100%) classic YWHAE-NUTM2high-grade endometrial stromal sarcomas and the 3 unusual high-grade endometrial stromal sarcomas which prompted FISH studies confirming YWHAErearrangement in 2 tumors. Genomic PCR confirmed the presence of BCORexon 16 internal tandem duplication in the third case. Diffuse BCOR staining was strong in three and weak in one BCOR-rearranged high-grade endometrial stromal sarcoma while absent in the remaining four BCOR-rearranged tumors. BCOR staining was weakly positive in <5% of tumor cells in 4 of 66 (6%) low-grade endometrial stromal sarcomas and 1 of 18 (6%) endometrial stromal nodules and weakly to moderately positive in <5–40% of tumor cells in 6 of 31 (19%) leiomyosarcomas. No BCOR staining was seen in the remaining low-grade endometrial stromal sarcomas, endometrial stromal nodules, leiomyosarcomas, or any of the leiomyomas. BCOR immunohistochemical staining is a highly sensitive marker for YWHAE-NUTM2high-grade endometrial stromal sarcoma with both classic and unusual morphology and identifies a subset of high-grade endometrial stromal sarcoma with BCORalterations, including BCORrearrangement and internal tandem duplication.

The distinction between undifferentiated melanoma (UM) or dedifferentiated melanoma (DM) from undifferentiated or unclassifiable sarcoma can be difficult and requires the careful correlation of clinical, pathologic, and genomic findings. In this study, we examined the utility of mutational signatures to identify patients with UM/DM with particular attention as to whether this distinction matters for treatment because the survival of patients with metastatic melanoma has dramatically improved with immunologic therapy, whereas durable responses are less frequent in sarcomas. We identified 19 cases of UM/DM that were initially reported as unclassified or undifferentiated malignant neoplasm or sarcoma and submitted for targeted next-generation sequencing analysis. These cases were confirmed as UM/DM by harboring melanoma driver mutations, UV signature, and high tumor mutation burden. One case of DM showed melanoma in situ. Meanwhile, 18 cases represented metastatic UM/DM. Eleven patients had a prior history of melanoma. Thirteen of 19 (68%) of the tumors were immunohistochemically completely negative for 4 melanocytic markers (S100, SOX10, HMB45, and MELAN-A). All cases harbored a dominant UV signature. Frequent driver mutations involved BRAF(26%), NRAS(32%), and NF1(42%). In contrast, the control cohort of undifferentiated pleomorphic sarcomas (UPS) of deep soft tissue exhibited a dominant aging signature in 46.6% (7/15) without evidence of UV signature. The median tumor mutation burden for DM/UM vs UPS was 31.5 vs 7.0 mutations/Mb (P< .001). A favorable response to immune checkpoint inhibitor therapy was observed in 66.6% (12/18) of patients with UM/DM. Eight patients exhibited a complete response and were alive with no evidence of disease at the last follow-up (median 45.5 months). Our findings support the usefulness of the UV signature in discriminating DM/UM vs UPS. Furthermore, we present evidence suggesting that patients with DM/UM and UV signatures can benefit from immune checkpoint inhibitor therapy.