Modern Pathology最新文献

HER2 gene amplification and heterogeneity influence anti-HER2 therapy efficacy, but quantitative assessment of HER2 expression heterogeneity remains scarce. This study evaluates HER2 protein heterogeneity and its effect on neoadjuvant therapy (NAT) outcomes in HER2-positive breast cancer (BC). In total, 295 HER2-positive BC patients treated with neoadjuvant anti-HER2 therapy were included. HER2 expression was quantified by the percentage of HER2-stained cells across intensities, and heterogeneity was assessed using quadratic entropy (QE). The overall pCR rate was 51.86%. Patients with HER2 heterogeneity showed a 24.5% pCR rate, significantly lower than the 62.5% in patients with HER2 homogeneity (P < 0.001). HER2 heterogeneity QE (HER2-HQE) correlated with the percentage of HER2 IHC 3+ tumor cells and hormone receptor (HR) status (P < 0.001). Residual cancer burden (RCB) was significantly associated with HR status, tumor stage, lymph node stage, HER2 IHC 3+ cell percentage, and HQE grade (P ≤ 0.017). No correlation was found with age, histological grade, or Ki67 index. In the HER2 IHC 3+ ≥95% subgroup, neither pCR rate nor RCB grade after NAT was associated with traditional clinicopathological parameters, even HER2-HQE and HR status (P≥0.154). AJCC anatomical stage significantly improved post-NAT for low/intermediate HER2-HQE subgroups (P < 0.001), but not for high HQE cases. In summary, HER2 heterogeneity and HR status are key predictors of anti-HER2 NAT outcomes. Higher HER2-HQE is associated with increased RCB. Quantifying HER2 heterogeneity via HQE and percentage of HER2 IHC 3+ may help optimize therapeutic strategies in clinical practice.

Light chain proximal tubulopathy (LCPT) is a rare kidney disorder associated with monoclonal gammopathy, traditionally defined by crystalline inclusions in proximal tubular epithelial cells. A poorly recognized variant, fibrillary LCPT, is characterized by large non-amyloid fibrillar aggregates but has remained inconsistently classified, leading to diagnostic uncertainty. We aimed to clarify its clinicopathologic features and refine its position within the LCPT spectrum. We retrospectively analyzed 41 biopsy-proven LCPT cases, reclassified by ultrastructural features into crystalline LCPT (n = 22), fibrillary LCPT (n = 11), amyloid LCPT (n = 2), and LCPT with lysosomal indigestion (n = 6). Fibrillary LCPT showed irregular cytoplasmic fibrils measuring 6-18 nm in diameter, arranged in intersecting, fishbone-like, or compact bundle-like patterns, confined to proximal tubular epithelial cells. These fibrils were Congo red-negative and not apparent on light microscopy. Conventional immunofluorescence on frozen tissue was uniformly negative in fibrillary LCPT, and pronase-digested paraffin immunofluorescence detected κ restriction in only 7/11 cases; immunoelectron microscopy confirmed κ light chain labeling in all paraffin IF-negative cases, yielding 100% diagnostic sensitivity. Clinically, most fibrillary LCPT cases were associated with monoclonal gammopathy of renal significance (MGRS, 72.7%), with the remainder linked to multiple myeloma (27.3%). This subtype was commonly associated with Fanconi syndrome (FS) and reduced estimated glomerular filtration rate (eGFR), with a subset of cases developing acute kidney injury. Following clone-directed therapy, a majority of patients achieved stable renal function and 85% showed improvement in tubular dysfunction, although proteinuria reduction was less pronounced than in crystalline LCPT. Fibrillary LCPT represents a distinct, non-amyloid entity characterized by Congo red-negative fibrils, κ light chain restriction, and frequent association with FS and MGRS. We propose a refined four-tier ultrastructural classification comprising four LCPT subtypes (crystalline LCPT, fibrillary LCPT, amyloid LCPT, and LCPT with lysosomal indigestion) to improve diagnostic precision and guide management.

Accurate assessment of visceral pleural invasion (VPI) is essential for staging and prognostication in non-small cell lung cancer (NSCLC), yet distinguishing elastin-rich pleural layers on routine hematoxylin and eosin (H&E) sections remains a diagnostic challenge. To overcome the cost and workflow delays associated with special elastic stains, we developed a deep learning pipeline that generates a virtual elastin stain, termed synthetic eosin-based elastin fluorescence (EBEF), directly from standard brightfield H&E slides. A key innovation of this study was the use of intrinsic eosin fluorescence from the same H&E section to create a perfectly co-registered, high-fidelity ground truth for training a conditional generative adversarial network, eliminating the spatial mismatches common in multi-slide approaches. In a multi-institutional validation, supplementing H&E review with synthetic EBEF significantly improved pathologists' diagnostic accuracy for VPI compared with H&E alone (P < 0.0001). Notably, the pre-analytical factors that optimized model performance, including thinner tissue sections (1-3 μm) and high-resolution scanning, also enhanced the perceptual contrast of elastin for pathologists, demonstrating a strong synergy between computational and conventional diagnostic optimization. This study establishes and validates a robust framework for high-fidelity virtual staining that improves diagnostic accuracy and provides a scalable pathway for integrating deep learning-based tools into routine digital pathology. The proposed approach offers a practical and cost-effective alternative to ancillary special stains in NSCLC evaluation.

The extent of residual disease after neoadjuvant chemotherapy (NAC) in patients with breast cancer (BC) holds prognostic value. However, current practices for reporting post-NAC BC specimens according to the ypTNM classification vary. This study aimed to map these practices and provide recommendations for standardization. A survey was developed and globally circulated to pathologists with a special interest in BC through personal networks and working group mailing lists. The survey included general questions about tumor diameter assessment, as well as graphical scenarios presenting different distributions of tumor cells. We did not provide definitions mentioned in reporting guidelines to capture unbiased current real-world practices. A total of 208 pathologists from 35 countries completed the survey. Almost all responding pathologists (97.1%) reported the ypTNM in daily practice. Despite self-reported strict adherence to the eighth edition of the international ypTNM classification, we found substantial variation in practice concerning the application of this staging system, particularly in cases with an uneven distribution of scattered residual disease. Notably, 57.2% of respondents reported measuring the largest "continuous cluster of tumor cells," but the interpretation of this definition varied widely. This international survey identifies the challenges and practice heterogeneity in the current application of the ypTNM staging system, which hampers the value of ypTNM reporting in daily practice. To enhance reproducibility and to provide more reliable post-NAC risk stratification, we recommend adopting standardized reporting with clearer pattern-based definitions of the ypTNM guidelines, supplemented with the elements of the residual cancer burden system.

Hepatoid lung carcinomas, similar to hepatoid carcinomas of other sites, are defined as extrahepatic tumors exhibiting divergent hepatocellular differentiation. Uniquely, hepatoid carcinomas of lung origin are reported to commonly express only HepPar1 - a hepatocellular marker, which recognizes mitochondrial enzyme carbamoyl-phosphate synthetase-1 (CPS1). Recently, HepPar1/CPS1 was found to accumulate in lung adenocarcinomas (LUAD) harboring STK11 mutations, presumably as a genotype-associated metabolic adaptation. The impact of these insights on the concept of hepatoid lung carcinoma has not been explored. Here, we performed a detailed clinicopathologic and genomic analysis on carcinomas prospectively regarded as hepatoid with isolated HepPar1 expression (n=17). We found that while robustly positive for HepPar1, these tumors were entirely negative for an extended panel of other hepatocellular markers (AFP, Arginase1, Glypican3, Albumin-ISH). Morphologically, tumors exhibited solid-trabecular architecture with expanded granular-vacuolated-clear cytoplasm, thus evoking hepatoid morphology; however, focal-to-moderate intracytoplasmic mucin was consistently present and hepatoid resemblance was variable. Pneumocytic markers (TTF1, Napsin A) were entirely negative (except for cytoplasmic TTF1), commonly leading to diagnostic challenges at metastatic sites. Remarkably, next-generation sequencing revealed invariable STK11 mutations/loss (P<0.00001 vs unselected LUAD, n>2.5K). Patient survival was dismal (median: 5.8 vs 25 months for stage-matched LUAD, P=0.0002). Tumors harbored high mitochondrial content by electron microscopy and other methods. For comparison, we reviewed conventional, predominantly acinar LUAD with HepPar1 expression (n=22), and found that they also lacked any other hepatocellular markers, had invariable STK11 mutations/loss, increased granular cytoplasm, lower TTF1 and poor prognosis. We conclude that isolated HepPar1 expression in LUAD reflects mitochondrial adaptation to STK11 mutations rather than bona fide hepatocellular differentiation, and that HepPar1-expressing solid and granular adenocarcinomas (SAGA) represent an undifferentiated (solid, TTF1-negative) variant in this spectrum of tumors. Recognition of these tumors is warranted due to their exceptionally aggressive behavior, distinct pathogenomic features and common association with diagnostic challenges.

A subset of perivascular epithelioid cell neoplasms (PEComas) is histologically malignant and at high risk for metastasis, and there is limited literature available on the genetic features of these lesions. In addition to driver alterations in the tuberous sclerosis complex/mammalian target of rapamycin pathway or TFE3 fusions, recurrent ATRX mutations have been identified in malignant PEComas. ATRX mutations have been tightly associated with the alternative lengthening of telomeres (ALT) phenotype in a variety of other tumor types. Whether malignant PEComas-regardless of ATRX mutational status-harbor the ALT phenotype has not been characterized. We conducted immunohistochemistry (IHC), next-generation sequencing (NGS), and telomere-specific fluorescence in situ hybridization (FISH) on a cohort of 32 malignant PEComas to evaluate for the ALT phenotype and to correlate with underlying genomic features. TSC1/2/mTOR/RICTOR mutations or TFE3 translocations were detected in 16 of 31 (52%) cases by NGS. Recurrent ATRX alterations were identified in 10 (32%) cases. Sixteen cases underwent ALT FISH, 8 of which harbored ATRX alterations by NGS and/or ATRX loss based on IHC, and 8 cases without detectable ATRX alterations by NGS or loss based on IHC. Twelve (75%) of these 16 cases demonstrated the ALT phenotype by FISH. All 8 (100%) cases with ATRX mutations were ALT positive by FISH. In 8 (50%) cases without ATRX alterations based on NGS, 4 cases demonstrated ALT by FISH. Of these 4 ALT-positive cases lacking ATRX genomic alterations, IHC revealed ATRX protein loss in 1 case. Overall, ATRX alterations were identified in 75% of ALT-positive tumors. Our study provides the first correlation between ALT and genomic features of malignant PEComas, demonstrating that ATRX alterations invariably predict ALT, but that a subset of tumors without mutations in known ALT suppressor genes also activate ALT. These findings confirm that the association between ATRX alterations and ALT observed in other tumor types applies to PEComas and indicates that in a subset of cases, ALT may be activated by ATRX-independent mechanisms.

Vascular anomalies encompass a heterogeneous group of vascular tumors and malformations and are most often caused by singular somatic variants that arise in utero. The presence of multiple variants, including those in at least 2 different genes (multigenic variants), is rare in these mosaic disorders, and their contribution to histology and phenotype has not been well characterized. We present our molecular and histologic experience with multiple variants in a large cohort of individuals with vascular anomalies and other suspected somatic mosaic disorders. We performed a retrospective review of 1299 individuals with suspected somatic mosaic disorders who were sequenced at our institution and identified 92 (7%) individuals with at least 2 clinically reportable variants. Our cohort includes 18 individuals with multigenic variants with unknown phenotypic implications and 13 individuals with well-described multigenic variants with clear phenotypic significance. For many individuals, the multiple variants had clear clinical implications, including variants that may respond to targeted therapy (n = 73) or germline variants (n = 33) with surveillance implications for that individual and their immediate relatives. Provided clinical information and available hematoxylin and eosin-stained slides (n = 37) were reviewed, and lesions were classified according to established guidelines whenever possible. The majority of lesions had histologic features concordant with the observed genetic variants. However, 4 out of 18 lesions with multigenic variants and unknown phenotypic implications had unique or unusual histologic features that did not fit into a specific diagnostic category. Our results demonstrate that broad sequencing of vascular anomalies, even in patients with known pathogenic variants, can be useful to identify additional clinically relevant variants that may refine diagnosis, respond to targeted therapy, or reveal a germline disorder that contributes to phenotype and may require increased surveillance.

Periductal stromal tumors (PDST) of the breast are rare biphasic neoplasms with morphologic similarities to phyllodes tumors (PTs). The histologic spectrum of PDST is broad but has not been well characterized, and its genetic underpinnings remain unknown. We profiled PDST by targeted next-generation sequencing in correlation with their histomorphology, immunophenotype, and clinical characteristics (n = 15). All patients were female, including 2 with Li-Fraumeni syndrome (LFS), with a mean age of 48 years. Forty-two percent had synchronous or metachronous PT and/or fibroadenoma. Most PDST expressed CD34 (15/15), smooth muscle actin (12/14), and at least focal nuclear HMGA2 (8/12) and/or beta-catenin (6/12). High-grade PDST (HGPDST, n = 8) were defined by marked nuclear pleomorphism (8/8), and most had high (≥10 mitoses/10 high-power field) and/or atypical mitotic activity (7/8) and pleomorphic multinucleated tumor cells (7/8). All HGPDST had p53 (88%, 7/8) and/or Rb/CDKN2A (75%, 6/8) alterations by next-generation sequencing or immunohistochemistry. p53 Aberrations correlated with the presence of pleomorphic multinucleated tumor cells. Both patients with LFS had HGPDST with loss of heterozygosity of the germline TP53 variant. Other altered genes in HGPDST included EGFR (25%, 2/8), NF1 (25%, 2/8), TERT promoter, PIK3CA, CDKN2A/B, LZTR1, KMT2B, and ARID2 (1 case each). Low-grade PDST, which lacked marked pleomorphism, high mitotic activity, or atypical mitoses (n = 7), had simpler genomes than HGPDST and lacked bona fide cancer gene alterations, with TERT promoter mutation in 1 case. Copy number alterations in PDST overlapped with those reported in PT, including 13q loss in all HGPDST. Copy number profiling revealed shared clonality of synchronous low-grade PDST and PT in 1 patient. In summary, we describe herein the genetic landscape of PDST, demonstrate correlation of genetic features with high-grade vs low-grade histology, and identify TP53 among key oncogenic drivers of HGPDST, including in LFS. The genetics of HGPDST overlap with borderline or malignant PT, consistent with their classification as PT variants that arise through a MED12-independent pathway.

Adenocarcinoma of the lung (LUAD) is common and highly lethal. Clinical grading of LUAD strongly predicts recurrence and survival after surgery and is determined by morphological assessment of histological growth patterns in resected tumors. In particular, the 2 archetypally lethal morphologies, solid, and micropapillary growth patterns, are highly distinct from each other, but little is known about their defining molecular features or how their appearances are related to their biology and mechanisms of lethality. Pure epithelial growth patterns and subregions were identified within 7 distinct LUAD growth patterns across 51 resected tumors and characterized using NanoString GeoMx digital spatial profiler in 160 epithelially pure regions of interest. Results were validated in 27 cases at the protein level using Akoya PhenoImager multiplex immunofluorescence, in 432 cases at the RNA level with TempO-Seq, and in 30 cases with an independent GeoMx digital spatial profiler. Analyses of gene expression reveal fundamental divergent evolutionary trajectories leading to solid and micropapillary growth. Additionally, we identify recurrent localized intratumoral plasticity in both growth patterns. These states can explain the origins of growth patterns and their mechanisms of virulence. Our work highlights dramatic divergence in gene expression programs between highly lethal modes of LUAD tumor growth. We go on to show how microscopically localized hypoxia in the primary tumor helps to establish and maintain cellular survival strategies and tumor architecture, suggesting morphology-specific mechanisms of LUAD tumor metastasis and suggesting new therapeutic vulnerabilities.

Follicular lymphoma (FL) patients have variable outcomes, underscoring the need for biomarkers for improved risk stratification. Current FL grading systems, based on subjective centroblast counts, suffer from poor reproducibility, despite evidence linking grade 3 FL to worse prognosis. We aimed to identify objective biomarkers for centroblasts and centrocytes to improve FL prognostication. We reanalyzed publicly available spatial and single-cell transcriptomic data from normal germinal centers (GCs) and FL samples. Reanalysis revealed distinct gene expression profiles: AICDA (AID) and CXCR4 highly expressed in GC dark zone cells (centroblasts), and CD40 and TFRC (CD71) in light zone cells (centrocytes). Single-cell RNA sequencing of FL samples further showed AID and CXCR4 overexpression in malignant Grade 3A cells and CD40 and CD71 in grade 1-2 cells. We validated these findings using immunohistochemistry (IHC) (single and multiplex) on tonsils and 59 FL specimens (42 Grade 1-2, 17 Grade 3). Grade 3 FL showed significantly higher expression of AID, CD71, and Ki67 compared to grade 1-2; with CXCR4 approaching significance. Receiver operating characteristic (ROC) curve analysis identified optimal cut-offs for AID (1.54%), CXCR4 (21.9%), Ki67 (21.6%), and CD71 (7.57%) to distinguish grade 3 from grade 1-2 FL, with AID showing the best discriminatory ability. Crucially, AID expression evaluation showed reproducibility across two different digital algorithms and two independent visual observers. Furthermore, we observed a trend toward shorter disease-specific survival (DSS) in patients with both FL grade 3 and high AID expression. This prognostic observation held true regardless of whether AID overexpression was assessed via digital evaluation (cut-off: 1.54%) or visual estimation (cut-off: 2%). In conclusion, AID, CXCR4, CD71, and Ki67 are promising biomarkers for objectively identifying FL grade 3, potentially enhancing the reproducibility of grading and serving as independent prognostic tools. Further clinical validation in uniformly treated FL cohorts is warranted.