Pathologica - Journal of the Italian Society of Anatomic Pathology and Diagnostic Cytopathology

Interventional pathologist and intraoperative surgical margin evaluation of radical prostatectomy specimens ex-vivo confocal microscopy vs. frozen section evaluation

Alessia Cimadamore, Liang Cheng, Antonio Lopez-Beltran, Rodolfo Montironi — Fri, 06 Feb 2026 00:00:00 +0000

Dear Editor,

We read with interest the contribution by Sofia Asioli et al. on Interventional Pathologists (IPs) and real-time histological evaluation in surgical and outpatient settings based on ex-vivo confocal microscopy (CFM) ¹. The authors review clinical applications in various fields, such as neurosurgery, dermatology and breast pathology, with some reference to prostate pathology, showing the benefits of such technology.

Our interest in the contribution by Asioli et al. is related to the adoption of CFM coupled with the involvement of IPs in the intraoperative evaluation of surgical margins of fresh radical prostatectomy specimens (RPSs). Prof. B. Rocco and his team ² introduced us to such a topic. It is called en-face margin evaluation: an entire surface area of the so-called capsule of the RPS is inspected with a CFM vs. the boundary between the prostate parenchyma and the so-called inked capsule at ~ 5 mm intervals in the conventional intraoperative frozen section analysis (IFS) (including the NeuroSAFE technique) ³.

Standardizing the pathologic assessment of HER2-Low and HER2-Ultralow breast cancer: Insights from a national Italian project

Fri, 06 Feb 2026 00:00:00 +0000

Objective

Novel therapies for HER2-(ultra)low breast cancer (BC) have changed the traditional binary classification of HER2 status into a multi-tiered system that captures the full spectrum of expression as determined by immunohistochemistry (IHC). Several pre-analytical and analytical variables can influence the accurate identification of HER2 expression within the lowest IHC range.

Herein, we present the findings of a national Italian project addressing the diagnostic challenges associated with HER2-(ultra)low BCs and their proper classification.

Material and Methods

A total of 121 pathologists from various regions and Institutions were recruited and asked to: i) complete an online survey addressing key pre-analytical and analytical issues affecting HER2 status reporting in BC; ii) score 8 cases of IHC HER2-low whole slide images (WSI) shared online; iii) participate in on-site meetings to discuss the results and evaluate 8 additional challenging WSIs. The assessments were subsequently compared to those of an expert panel.

Results

Several pre-analytical and analytical concerns emerged from the questionnaire, like reporting resection cold ischemia time and decalcification agent choice and the employment of proper HER2 controls. Regarding the WSIs, a substantial overall agreement (69%) with the expert panel was observed (74% in the online phase and 64% in the on-site phase), along with substantial to excellent consensus (≥ 61%) in over 60% of the samples. Most discordances emerged in the on- site cohort, which was enriched with challenging cases, particularly within the HER2 0+/ultralow spectrum, which demonstrated an overall agreement of 48%.

Conclusions

This nationwide study highlights the complexities of accurately classifying HER2-(ultra)low breast cancers, particularly within the HER2 ultralow subset. While a substantial level of agreement with expert assessments was achieved, the variability observed in more challenging cases underscore the need for standardized interpretation criteria, enhanced training, and continuous quality assurance measures.

PIK3CA testing in HR+/HER2− metastatic breast cancer: assessing pathology laboratories capacity and needs

Fri, 06 Feb 2026 00:00:00 +0000

The management of hormone receptor-positive/HER2-negative (HR+/HER2−) metastatic breast cancer (MBC) relies on molecular testing to inform treatment decisions. PIK3CA mutations, present in ~40% of cases, represent a key predictive biomarker for PI3K-pathway–targeted therapies. Despite its clinical relevance, PIK3CA testing continues to face challenges related to laboratory organization, standardization, and access. We conducted a nationwide, cross-sectional survey to evaluate current practices and institutional readiness for PIK3CA testing in Italy, in the context of the anticipated expansion of PI3K-targeted therapies, including inavolisib. A total of 118 healthcare professionals from institutions across 15 regions participated, providing data on test availability, laboratory workflows, analytical methodologies, accreditation status, and implementation barriers. Descriptive statistics were used for analysis. Overall, 88.1% of institutions reported the ability to perform PIK3CA testing, with 57.6% offering on-site analysis. Testing was predominantly performed in pathology laboratories (76.5%), followed by molecular biology (16.2%) and genetics laboratories (7.4%). However, 46.6% of institutions lacked formal molecular accreditation, and ISO:15189 certification remained uncommon. Pre-analytical workflows relied mainly on formalin-fixed paraffin-embedded (FFPE) tissue samples (89.7%), with limited routine use of liquid biopsy. Next-generation sequencing (NGS) was the most frequently adopted analytical approach (45.6%), followed by combined NGS and PCR-based strategies (36.8%). Most institutions reported turnaround times of 7–15 days. In conclusion, this updated survey indicates progress in access to PIK3CA testing and consolidation of NGS-based methodologies in Italy. Nevertheless, persistent gaps in accreditation, heterogeneous workflows, and limited integration of liquid biopsy highlight ongoing challenges in standardization and diagnostic equity. Coordinated national strategies will be essential to ensure consistent, high-quality molecular diagnostics in HR+/HER2− MBC.

Distinction of thymic carcinoma and type B3 thymoma using ancillary biomarkers

Fri, 06 Feb 2026 00:00:00 +0000

Objective. Thymic carcinomas (TC) are rare and understudied tumors. Pitfalls exist with TC diagnosis, and biomarkers are needed to support the pathologist. Here, we tested a series of biomarkers to differentiate TC from type B3 thymoma.

Methods. A consecutive series of 48 patients, 26 with TC and 22 with type B3 thymoma entered the study. Immunohistochemical expression of CD5, CD117, BAP1, MTAP, Ki-67 was evaluated. CDKN2A status was assessed by FISH.

Results. CD5 and CD117 were expressed in TC only (n = 19 and n = 20 respectively). Five TC did not show CD5 or CD117 expression. BAP1 expression was lost in 3 TC, while MTAP staining was absent in 3 TC and 1 type B3 thymoma. CDKN2A deletion was observed in 4 TC and 1 type B3 thymoma. CD5 and CD117 showed a perfect specificity for TC and a good sensitivity, especially when combined (0.81). The addition of the other markers improved the sensitivity (0.85) with a slight decrease in specificity (0.95). Indeed, one type B3 thymoma harboured CDKN2A deletion with MTAP loss of expression.

Conclusions. CD5 and CD117 are the best markers for TC. While the addition of other markers (i.e., BAP1 loss, MTAP loss and CDKN2A deletion) might be useful in cases negative for CD5 and CD117, rare cases of type B3 thymoma might harbor these alterations.

Immunohistochemical and molecular profiling of uveal melanoma: clinicopathological correlations from an Italian cohort

Fri, 06 Feb 2026 00:00:00 +0000

Objective. Uveal melanoma (UM) is the most common primary intraocular malignancy in adults, characterized by distinct histopathological and molecular features and often associated with poor prognosis due to its high metastatic potential. While histology, BAP1 status, and chromosomal changes are established prognostic markers, integration of morphological, immunophenotypic, and molecular data is evolving.

Methods. We retrospectively analyzed 84 UM cases from a single institution using an integrated approach combining histological classification, immunohistochemical profiling, and targeted next-generation sequencing with a 63-gene panel. Tissue microarrays were used for immunophenotyping, and mutation data were stratified by prognostic outcomes.

Results. Most tumors were localized to the choroid and predominantly exhibited spindle-cell morphology. Mutations in GNAQ or GNA11 were identified in 83% of sequenced cases. Loss of BAP1 expression correlated with epithelioid histology and denser T-cell infiltration yet lacked PD-L1 expression. Aberrant p53 staining was more frequent in spindle-cell tumors, though TP53 mutations were rare, suggesting functional inactivation through other mechanisms. Notably, mutations typically associated with cutaneous melanomas (e.g., BRAF, KIT, CDKN2A) were also detected, particularly in a single iris melanoma, suggesting site-specific molecular convergence. Additional recurrent alterations were found in NOTCH1, PTEN, PIK3CA, and KDR, implicating the mTOR and VEGF signaling pathways. A high mutational burden, along with mutations in genes such as H3F3A, IDH2, JAK3, and ESR1, was more frequent in tumors with poorer prognosis, supporting their potential role in disease aggressiveness.

Conclusions. This study highlights the heterogeneous molecular landscape of UM and underscores the importance of integrating histopathological and molecular data for improved prognostic stratification. The identification of potential therapeutic targets and atypical mutations typically associated with other melanoma subtypes suggests avenues for future research and tailored therapeutic strategies.

The clinical impact of precise assessment of predictive biomarkers in gastroesophageal cancer: focus on the PD-L1 combined positive score (CPS) and tumor area positivity (TAP) systems

Fri, 06 Feb 2026 00:00:00 +0000

Accurate assessment of PD-L1 expression is crucial for therapeutic decision-making in esophageal, esophago-gastric junction, and gastric cancers, where immune checkpoint inhibitors have become integral to first-line treatment in selected patients. This review provides an updated, practice-oriented summary on PD-L1 immunohistochemistry evaluation, with emphasis on the emerging Tumor Area Positivity (TAP) scoring system together with established Combined Positive Score (CPS) and Tumor Proportion Score. First, we examine the clinical relevance and use in clinical trials of each scoring method, and the pre-analytical and analytical variables influencing PD-L1 interpretation. Then, we address advantages and disadvantages of each scoring system, including a thorough analysis and pictorial interpretation guide of the recently introduced TAP score. Indeed, thanks to a visual-estimation-based assessment of PD-L1 expression, TAP has improved reproducibility and reduced scoring time, but large-scale validation is ongoing and certain interpretive challenges remain. Finally, we propose a standardized reporting template to enhance consistency in diagnostic practice, together with our perspective on future improvements and challenges of PD-L1 assessment.

Next-generation sequencing methodologies to identify patients for targeted therapy: focus on HR+/HER2− metastatic breast cancer

Fri, 06 Feb 2026 00:00:00 +0000

Alterations in the phosphoinositide 3-kinase (PI3K)/AKT/PTEN signaling pathway are a well-recognized mechanism of resistance in hormone receptor-positive, HER2-negative metastatic breast cancer (HR+/HER2- mBC). These alterations are present in approximately half of patients with HR+/HER2- mBC. The major alterations in the pathway are somatic mutations in the PIK3CA (40–45%) and AKT1 (5%) genes, and loss-of-function alterations in PTEN (5–10%). New targeted agents that act against these alterations have been developed. Therefore, it is important to determine the mutational status of genes in this pathway to potentially offer a therapeutic alternative for these patients. In this review, we discuss the clinical and biological significance of PI3K pathway alterations in HR+/HER2− mBC, focusing on tumors that progress following endocrine therapy and CDK4/6 inhibitor treatment. We then highlight how different diagnostic strategies, including sample type, testing methodology, and timing, can improve the identification of patients who are eligible for targeted therapies and promote the effective integration of molecular diagnostics into routine clinical care.

Relevance of next-generation sequencing in the differential diagnosis of meningeal mesenchymal tumors: primary meningeal dedifferentiated chondrosarcoma of the cavernous sinus

Nicolò Caldonazzi, Gonzalo Hernandez Gamero, Andrea Mafficini, Valeria Barresi — Fri, 06 Feb 2026 00:00:00 +0000

Primary mesenchymal tumors of the meninges include various types, with meningioma being the most prevalent. Non-meningothelial primary tumors of the meninges are uncommon and present diagnostic difficulties due to their potential morphological similarities with each other and with the anaplastic subtype of meningioma. Intracranial dedifferentiated chondrosarcoma is extremely rare and is defined by the presence of both a conventional chondrosarcoma component and a non-cartilaginous sarcoma component. Diagnosing this tumor can be particularly challenging in biopsy samples or when the chondrosarcoma component is not prominently represented. We report a rare case of dedifferentiated chondrosarcoma of the cavernous sinus in a specimen containing only the dedifferentiated component and exhibiting an IDH1 mutation, underscoring the importance of genetic characterization for the differential diagnosis of tumors in this anatomical region. Given the recent evidence supporting the efficacy of IDH inhibitors in chondrosarcomas, identifying mutations in these genes may also have significant therapeutic implications.

Gastric perineurioma detected in routine endoscopy biopsy practice: case report and review of clinico-pathological findings in the literature

Fri, 06 Feb 2026 00:00:00 +0000

Perineuriomas are soft tissue neoplasms composed almost entirely of cells resembling perineurium and their occurrence in gastrointestinal tract is quite rare, with most of reported cases occurring in the colon. No more than 12 cases of gastric perineurioma have been reported to date, and the diagnosis in a routine endoscopy biopsy service can be challenging given the broad differentials of a bland-looking spindle cell proliferation in gastric mucosa. Here we present a case of a gastric perineurioma in a 39-year-old woman detected in routine endoscopy biopsy where a high degree of suspicion of the pathologist and targeted immunohistochemical investigations for perineurial markers EMA and GLUT1 allowed the correct diagnosis. Reviewing the literature, it appears that gastric perienuriomas most commonly occur in mid-age females and are often detected in routine endoscopies, with appearance of a benign-looking mid-sized sessile polyp. Special attention is needed to discriminate with GIST to avoid inappropriate further investigations and overtreatment. Moreover, we speculate that these lesions are likely underrecognized in busy routine gastrointestinal biopsy service, as most are probably dismissed as benign mesenchymal polyps/submucosal proliferations after exclusion of GIST or are misdiagnosed as inflammatory fibroid polyp in the case of CD34 positivity. However, the case presented demonstrates that, in light of the apparently typical occurrence in mid-aged females with endoscopic impression of benign polyp of the body, general pathologists of routine biopsy services should also have a high degree of suspicion and maybe add a perineural marker to their panel.

Marked hepatic fibrosis with progression towards cirrhosis in generalized arterial calcification of infancy: an unreported association observed in a case carryng a novel ENPP1 variant

Fri, 06 Feb 2026 00:00:00 +0000

Generalized arterial calcification of infancy (GACI) is a rare autosomal recessive disorder characterized by dysregulated calcium-phosphate metabolism, leading to mineral deposition within the internal elastic lamina of medium- and large-sized arteries. This results in arterial wall thickening and luminal narrowing due to intimal hyperplasia, causing significant vascular disruption. Approximately 70% of cases (GACI type 1) are caused by biallelic loss-of-function mutations in the ENPP1 gene, with nearly 40 pathogenic variants reported. We report a case of an infant diagnosed with GACI type 1 who died at 7 weeks of age. The patient was delivered via cesarean section at 36 weeks of gestation after a pregnancy complicated by polyhydramnios. The parents were second-degree cousins, with a history of two neonatal deaths of unknown etiology and one miscarriage. Autopsy revealed diffuse arterial calcification with prominent involvement of the coronary arteries. Notably, the liver showed fibrosis progressing to cirrhosis. Genetic analysis through trio exome sequencing identified a novel homozygous nonsense variant in ENPP1 (c.553C > T; p.Gln185Ter), inherited from both parents. This stop-gain variant is predicted to produce a severely truncated, non-functional or absent protein. This case is notable for two key aspects: a previously unreported association between GACI and progressive hepatic fibrosis evolving into cirrhosis, and the identification of a novel pathogenic ENPP1 variant not previously described in the literature.