Since their initial description in 1948, Spitz tumours have always been a challenge in the field of dermatopathology and paediatric pathology. Advances in molecular pathology have confirmed they are associated with specific anomalies, mainly gene fusions. They display a wide range of clinical presentations and histological subtypes. Most cases are Spitz nevi and very few lesions match the criteria to be diagnosed as atypical Spitz tumours. Even fewer are labelled as Spitz melanomas. Follow-up studies of genetically characterized cases have repeatedly confirmed that, even if the regional lymph node is involved, the overall outcome remains favourable. The aims of this review are to cover the variety of morphological presentations of Spitz tumours and illustrate the most rare subtypes. When possible, we have pointed out the potential trends between some unusual morphological features and the frequently associated genetic drivers. Spitz tumours have many differential diagnoses, the main being superficial spreading melanoma, with overlapping morphological features in early lesions. Essential clues to discriminate Spitz from mimickers have been listed and illustrated.

The advent of "omics" technologies for high-depth tumor profiling has provided new information regarding cancer heterogeneity. However, a bulk omics profile can only partially reproduce tumor complexity, and it does not meet the preferences of pathologists used to perform an in situ assessment of marker expression, for instance, with immunohistochemistry. The NanoString GeoMx® Digital Spatial Profiler (DSP) is a platform for morphology-guided multiplex profiling of tissue slides, which allows the digital quantification of target analytes in different neoplastic settings. To illustrate the feasibility and opportunities offered by DSP from a pathologist's perspective, we applied DSP in three different representative neoplastic settings: breast carcinoma, thyroid anaplastic carcinoma, and biphasic mesothelioma. Because of the perfect overlap between the hematoxylin-eosin-stained slide and the GeoMx areas of interest, in breast carcinoma, two different antibodies allowed the distinction of the tumor cells from the surrounding tumor microenvironment. In biphasic mesothelioma, we could distinguish the epithelioid from the sarcomatoid neoplastic component, and in the thyroid, we easily separated the anaplastic areas from the well-differentiated carcinoma. DSP is a promising tool that combines traditional histological evaluation, allowing spatial assessment of a tumor and its surroundings, and innovative in situ digital profiling. Pathologists should not miss the opportunity to combine morphological and genomic analyses and be at the forefront of investigating the progression of dysplasia/neoplasia, low-grade or high-grade, epithelial/mesenchymal, and, more in general, overcoming the concept of in situ vs. bulk genomic methods.

Ki-67 index (Ki-67i) is integral to the grading of many tumours. There remains considerable variability across pathologists in methods used to determine Ki-67i and in their results. Manual counting (or "eyeballing") is widely used, but digital pathology tools such as web-based image analysis and artificial intelligence-assisted cell detection software have become available in daily pathology practice. This study aims to compare the accuracy and efficiency of manual and two digital methods of Ki-67i determination. H&E and Ki-67 immunohistochemical (IHC) slides/images of 12 gastrointestinal neuroendocrine tumours (GI-NETs) were provided to 8 pathologists to evaluate Ki-67i via manual estimation (ME; the "past"), web-based image analysis using cellular segmentation (AI4Path.ca; the "present"), and software-based image analysis with built-in AI algorithms (QuPath; the "future"). Data collected include Ki67i, time expended, total cells counted, and pathologists' confidence level in the reported result. Deviation of Ki-67i from a gold standard result (GS) was analyzed using multiple linear regression, and results were compared via paired t test. Our results found no statistically significant differences in Ki-67i deviation from GS when comparing ME and AI4P methods for all 12 cases. The QP Ki-67i detection accuracy varied significantly. ME was the method with the least time expenditure. Junior pathologists are less confident in ME. Grading consensus was comparable among all three methods. These findings suggest that while digital pathology can confer increased Ki-67i accuracy in some cases of GI-NETs, higher time expenditure and proper hotspot selection may represent barriers to the adoption of digital pathology methods in the future.

In autopsy practice, the thickness of ventricular walls is one of the parameters used to identify cardiac hypertrophy. The presented study aimed to compare ante- and postmortem measurements of ventricular wall thickness, (i) to determine a postmortem standardized localization and dissection method for ventricular wall measurements, and (ii) to determine the ability of postmortem measurements in recognition of antemortem hypertrophy. A single-center prospective study was conducted at the Institute of Legal Medicine in Hamburg, Germany. Sixty hearts were dissected alternating by the inflow-outflow or short-axis method, and the ventricular walls were measured at different locations and compared with the echocardiographic values of the end-diastolic phase during life of these individuals. The results showed measurement differences between the autoptic and echocardiographic values-for the left ventricle between 3.3 and 5.2 mm, for the right ventricle between 0.2 and 1.1 mm, and for the septum between 1.3 and 1.4 mm. Diagnostic performance of recognizing antemortem hypertrophy with postmortem measurement was poor, except for measuring the right ventricle and septum with the short-axis method (area under the ROC curve of 0.72 and 0.82, respectively). According to the results, cardiac changes may occur postmortem and need to be considered when used for diagnosing cardiac pathology. The postmortem diagnosis of left or right ventricular hypertrophy should always be made in conjunction with other, particularly cardiac, autopsy findings. An autoptic diagnosis of hypertrophy solely by a ventricular wall thickness > 15 mm or > 5 mm alone is not sufficient.

This review article focuses on the various primary osseous tumors of the orbit. Due to overlapping clinical, radiologic, and histologic features, differentiating these entities can pose significant challenges diagnostically. In this review, emphasis is placed on key distinguishing clinical, morphologic, immunophenotypic, and molecular characteristics. Also described are important prognostic details, recurrence risks, and the gold standard treatment methods for each entity. Relevant genetic syndrome associations are additionally covered. Orbital bone entities discussed include osteoma, osteoid osteoma, osteoblastoma, ossifying fibroma, fibrous dysplasia, aneurysmal bone cyst, osteosarcoma, Ewing sarcoma, and mesenchymal chondrosarcoma.

Sclerosing mucoepidermoid carcinoma (SMEC) of the salivary glands is a rare variant of low-grade mucoepidermoid carcinoma with scanty cellular atypia characterized by marked fibrosis/sclerosis and a rich inflammatory infiltrate. Herein, we report 25 unpublished cases of SMEC, two of them with prominent eosinophilia (2/25; 8%) and three with abundant IgG4-positive plasma cells (3/25; 12%). In our series of salivary SMEC, molecular analysis using fluorescence in situ hybridization (FISH) and/or next-generation sequencing (NGS) provided evidence of MAML2 gene rearrangement in 18 cases of the 21 analyzable cases tested (86%), while this gene locus was intact in 3 cases (14%). This study focuses on the diagnostic criteria of salivary SMEC given its challenge of abundant collagenous stroma, minimal residual neoplastic areas, and inconspicuous mucous cells. Follow-up data of our cases indicate that salivary SMECs have favorable outcomes. Molecular analysis for MAML2 gene rearrangement suggests that SMECs of salivary glands represent a rare variant of conventional low-grade MECs of salivary glands. In contrast, SMECs of the thyroid gland are genetically distinct from salivary-type thyroid MECs.

The majority of mastocytosis cases are characterized by an activating mutation in the KIT gene in codon 816. The detection of this alteration is of importance for proper diagnostic workup. Therefore, reliable and sensitive methods for the detection of KIT Codon 816 hotspot mutations in various types of patient samples are required. Since mutated cancer genes are often overexpressed, we evaluated the feasibility and sensitivity of KIT p.D816V detection by analysing mRNA/cDNA instead of genomic DNA. From 80 bone marrow trephines harboring a KIT p.D816 mutation, seven were only mutated by mRNA/cDNA pyrosequencing and 11 only by digital PCR analysis of genomic DNA. These results clearly demonstrate that detection of clinically relevant mutations in mRNA extracted from routinely processed decalcified archival bone marrow trephines is not only possible in a reliable fashion but under many circumstances advantageous. This enables the direct correlation of genomic data with high-quality morphological evaluation.

Succinate dehydrogenase (SDH)-deficient renal cell carcinoma (RCC) is a rare subtype of renal neoplasm predominantly affecting younger individuals. It is characterized by germline mutations in SDHx genes, particularly type B. Histologically, SDH-deficient RCC features eosinophilic cytoplasmic cells forming solid nests or microcysts, sometimes entrapping normal tubules. We present three SDH-deficient RCC cases with overlapping morphological features with fumarate hydratase-deficient RCC and TFEB-rearranged RCC, an appearance that has not been previously described. All tumors lacked SDHB expression and harbored pathogenic SDHB mutations, with the germline nature confirmed in two cases. Metastasis developed in two patients. Our case set highlights the diagnostic challenges of molecularly defined renal tumors and expands the morphological spectrum of SDH-deficient RCC with unusual histological features. Clinically, these tumors appear to be aggressive.

Fibroblastic and myofibroblastic neoplasms represent about 12% of pediatric soft tissue tumors. Most of these neoplasms in children are either benign or locally aggressive with rare metastasis, while malignant cases are uncommon. Diagnosing these tumors is challenging due to overlapping morphologies and the limited utility of immunohistochemistry. Advances in molecular techniques, especially RNA sequencing, have improved our understanding of the molecular drivers of these tumors, leading to better classification. Key molecular alterations, such as RTK and MAPK activation, are central in the development of tumors like infantile fibrosarcoma (IFS) and inflammatory myofibroblastic tumors (IMT). The identification of alternative fusions in IFS and IMT underscores the importance of an integrated diagnostic approach. Furthermore, new RTK-driven lesions, now included in the WHO's "NTRK-rearranged mesenchymal neoplasms", have been identified. This review provides an update on recent findings in RTK-driven myofibroblastic tumors and highlights novel entities still in need of classification.

The classification of TFEB-altered renal cell carcinoma (RCC) has been revised to include TFEB-rearranged RCC and TFEB-amplified RCC in the 2022 World Health Organization (WHO) Classification of Tumors of the Urinary System. Given the wide spectrum of TFEB-altered RCC in terms of morphology and clinical behavior, an accurate diagnosis is challenging yet crucial, particularly in aggressive cases. Moreover, the concurrence of TFEB gene rearrangement and amplification/gene copy number (GCN) gains was also observed, but there was limited knowledge of these cases. We presented three TFEB-rearranged RCC cases, one TFEB-amplified RCC case, and one case of concomitant TFEB-rearranged and -amplified RCC, comparing the similarities and differences among these three subgroups. Furthermore, we summarized the clinicopathological and molecular features of TFEB-rearranged RCC concomitant with TFEB amplification/GCN gains from the literature and the present study. TFEB-altered RCCs exhibit significant heterogeneity in morphology and clinical behavior while displaying similar immunohistochemical profiles, including positive staining for Melan-A, PAX8, and CD117, and negative staining for CK7. A typical biphasic "rosette-like" morphology has been observed in a proportion of TFEB-rearranged RCC concomitant with TFEB amplification/GCN gains, which has been noted in TFEB-rearranged RCC, but not in cases with only TFEB amplification. Notably, TFEB-rearranged RCCs concomitant with TFEB amplification/GCN gains tend to be aggressive, in contrast to the often indolent nature of TFEB-rearranged cases, irrespective of the extent of TFEB gene copy increase. Therefore, a TFEB FISH assay is essential for unclassified RCC cases that exhibit melanocytic marker expression, and fluorescent signals should be counted and interpreted acurrately.

The management of thyroid nodules is guided by the cytological classification provided by The Bethesda System for Reporting Thyroid Cytology. Notably, the biology of thyroid tumors in pediatric patients differs from that in adults, and there is limited research focused on pediatric cases. This study aimed to assess the effectiveness of the Bethesda system in pediatric patients treated at the largest tertiary pediatric thyroid center in Poland between 2015 and 2023. A retrospective analysis was conducted on 566 patients with thyroid nodules, of whom 555 underwent fine-needle aspiration biopsy (FNAB). A total of 217 patients underwent thyroid surgery. Of these, 206 had previously undergone FNAB with cytological evaluation at our center, while 11 patients underwent thyroid surgery due to a RET mutation or the need for an extended procedure. The initial FNAB results showed distribution across Bethesda categories as follows: 7.6% for category I, 54.6% for category II, 20.9% for category III, 4.1% for category IV, 7.6% for category V, and 5.6% for category VI. Among patients who underwent surgery, the distribution of Bethesda categories I through VI was 2.9%, 25.2%, 29.1%, 8.3%, 19.4%, and 15%, respectively. The risk of malignancy (ROM) from the initial FNAB was estimated at 33.3%, 11.5%, 22.2%, 4.8%, 84.4%, and 96.8% for Bethesda categories I through VI, respectively. In patients with autoimmune thyroiditis (AIT), the ROM was higher than in non-AIT patients for Bethesda categories I through IV, while it was lower in category VI. The sensitivity for detecting non-benign neoplasms across Bethesda categories III through VI was approximately 86% in both AIT and non-AIT patients. However, for papillary thyroid carcinoma, sensitivity in Bethesda categories V and VI was 86% in non-AIT patients but decreased to 61.5% in AIT patients. These findings emphasize the importance of considering surgical intervention in pediatric patients with Bethesda III-VI cytology, particularly in those with AIT.

Architectural distortion and basal plasmacytosis are the most widely recognized histologic features of chronic ileal inflammation. However, these features might be difficult to assess in small, poorly oriented, or superficial biopsies. Additional features of chronic mucosal damage, including pseudopyloric or pseudofoveolar metaplasia and Paneth cell hyperplasia, have been less commonly reported, and their broader appreciation could facilitate the diagnosis of chronic ileal inflammatory conditions. The prevalence of gastric-like (pseudopyloric and pseudofoveolar) metaplasia and Paneth cell hyperplasia was evaluated in 102 ileal biopsies obtained from patients with Crohn's disease (n = 47), ulcerative colitis with endoscopically normal ileum (n = 20) or with backwash ileitis (n = 20), and nonsteroidal anti-inflammatory drugs- (NSAIDs-) induced ileitis (n = 15). Gastric-like metaplasia was identified in 23% of CD and 13% of NSAID-induced ileitis cases, whereas it was absent among all ulcerative colitis cases. Pseudopyloric metaplasia, pseudofoveolar metaplasia, or a combination of both was documented in 13%, 2%, and 9% of Crohn's disease cases, respectively. NSAID-associated cases showed only pseudopyloric metaplasia. Paneth cell hyperplasia was detected in 43% of Crohn's disease cases, 13% of NSAID-induced ileitis cases, and 5% of backwash ileitis cases. Accordingly, pseudofoveolar metaplasia, pseudopyloric metaplasia, and Paneth cell hyperplasia are not uncommon in conditions causing chronic ileal inflammation. They are most frequently detected in Crohn's disease, but may also be present in NSAID-induced ileitis, whereas they are significantly less common in backwash ileitis and absent in normal ileum. Given the surface localization of pseudofoveolar metaplasia, its identification can be particularly helpful when dealing with poorly oriented or superficial samples.

Several distinctive round cell sarcomas have emerged by leveraging new testing modalities to include immunohistochemistry, next-generation sequencing, methylation array, and others. While Ewing sarcoma has led the way as the prototypic round cell sarcoma, more recently described round cell sarcomas of bone and soft tissue are now recognized which have unique clinical, morphologic, immunophenotypic, and genetic signatures. While each of these entities is less common than Ewing sarcoma, it is important to distinguish these tumors for correct diagnosis, prognostication, and potential treatment management. The focus of this review will cover CIC-rearranged sarcoma, BCOR-altered sarcomas, and EWSR1-non-ETS sarcomas to include recent developments in desmoplastic small round cell tumor as well as sarcomas with EWSR1/FUS::NFATc2 and EWSR1::PATZ1 gene fusions, highlighting the clinical, morphologic, and immunophenotypic clues to the diagnosis with recognition of each molecular diagnostic hallmark.

Correctly diagnosing neuroendocrine neoplasm (NEN) has become increasingly challenging, given that more histomorphologic and immunophenotypic NEN mimics have been identified in recent years. A systemic review was conducted on the 4795 consult cases submitted with initial diagnoses of NEN to a national reference center in China from 2013 to 2021. Among them, 443 cases were misdiagnosed as epithelial NENs after reevaluation with the help of immunohistochemical and/or molecular tests, ranging from 7.1 to 13.2%, with yearly increases. The misdiagnoses varied among age groups and tumor sites. Exocrine carcinoma was the most common (63.2%), followed by mesenchymal tumors. Other common tumors that were misdiagnosed included hepatocellular carcinoma, salivary gland tumor, and gastrointestinal stromal tumor. Aberrant expression of neuroendocrine markers was frequent (218/408, 53.4%), with diffuse positivity ranging from 8.2 to 51.7% for synaptophysin, chromogranin A, and INSM1 stains in all non-NEN cases. Selecting appropriate immunohistochemical stains based on H&E morphology is the key to avoiding diagnostic pitfalls. Medical history and molecular genomic information greatly assist in correctly diagnosing NENs and their mimics.