Genetic tests for lung cancer, such as the Oncomine Dx Target Test (ODxTT), are essential for patient management, and require RNA and DNA of sufficient quality and quantity from formalin-fixed paraffin-embedded (FFPE) samples. ODxTT fails due to poor nucleic acid preservation. Although enhancing the penetration of formalin into tissues improves nucleic acid preservation, the clinical utility of negative-pressure fixation methods remains unclear. Therefore, we investigated whether the negative-pressure fixation method using a vacuum fixation device improves ODxTT success rates and the quality of nucleic acids in FFPE samples from lung and other cancers, and examined the underlying mechanisms. The ODxTT success rate was higher with the negative-pressure fixation method than with the conventional method (96.5% vs. 81.2%), accompanied by increased RNA quality and yield; however, no marked differences were observed in DNA. RNA deteriorated more rapidly than DNA in resected mouse liver tissues, indicating that RNA is more labile than DNA during cold ischemia. Furthermore, in FFPE samples from colon, pancreatic, and prostate cancers, negative-pressure fixation consistently improved RNA DV200. These results indicate that the negative-pressure fixation method improves ODxTT success by accelerating formalin penetration, which may suppress RNase-mediated RNA degradation and preserve RNA quality.

This study clarifies the distinct clinicopathological and molecular features of amphicrine pancreatic carcinoma (AmpCA) through a case analysis and literature review. We present a case of AmpCA in a 73-year-old female, characterized by co-expression of acinar cell markers and neuroendocrine markers within individual tumor cells. Molecular profiling revealed high-risk mutations in KRAS G12V, TP53, and RB1, along with a high Ki-67 index (85%). Literature comparison confirms that AmpCA exhibits a unique bidirectional, single-cell differentiation pattern, fundamentally distinguishing it from the bicellular composition of mixed neuroendocrine-non-neuroendocrine neoplasms (MiNEN). We conclude that AmpCA is a distinct malignant entity requiring specific diagnostic criteria, and its molecular profile suggests potential for targeted therapies.

Undifferentiated carcinoma (UC) is a rare, highly aggressive subtype of pancreatic cancer; UC with rhabdoid features (UCRF) is particularly associated with poor prognosis. This study aimed to compare clinicopathologic characteristics and outcomes of UC between groups of patients defined by presence versus absence of RF. We retrospectively reviewed patients diagnosed with UC by EUS-guided tissue acquisition from 2017 to 2024. Histologically, 11 patients were classified as anaplastic UC and two had UC with osteoclast-like giant cell. Six patients had UCRF. Only 2 patients underwent surgery, while 8 received chemotherapy; 5 of these were treated with gemcitabine plus nab-paclitaxel. One patient (17%) in the UCRF group was INI-1-negative. CK7 was positive in 3 UCRF patients (50%) and in all 7 UC patients without RF. In the overall cohort, median OS showed no statistically significant difference between UCRF and UC without RF (3.9 vs. 12.9 months; log-rank p = 0.188). However, after excluding two patients with UCOGC, Kaplan-Meier analysis demonstrated a significantly shorter OS in UCRF (p = 0.009). These findings suggest that RF represents a potential adverse prognostic factor in pancreatic UC. Further studies are warranted to clarify biologic significance and treatment implications of RF in pancreatic UC.

Ancillary assays, including BAP1 (BRCA-1-associated protein 1), MTAP (methylthioadenosine phosphorylase), and Merlin immunohistochemistry (IHC), are effective in discriminating pleural mesothelioma (PM) from reactive mesothelial proliferations (RMP). IHC of Merlin, a protein product of the neurofibromin 2 gene, was most recently developed. Our previous study suggested uneven occurrence of Merlin loss within PM tissues. This was further investigated here in a total of 41 tissues of PM obtained from seven institutions. Partial or diffuse Merlin loss was detected with excellent interobserver agreement (kappa: 0.82). Of the 34 PM cases with interobserver consensus, partial and diffuse loss of Merlin expression were each observed in 13 cases (38%). PM with partial Merlin loss exhibited either discrete or intermingled spatial distribution of Merlin-lost cells. BAP1 or MTAP showed partial loss in none or only 5% of PM cases, respectively. The diagnostic sensitivity for PM was enhanced to 91% with addition of diffuse and partial Merlin loss to a combination of BAP1 and MTAP loss while maintaining 100% specificity. Although detection of Merlin loss as well as BAP1 and MTAP losses is effective for distinguishing PM from RMP, pathologists should be familiar with partial Merlin loss when interpreting Merlin IHC results.

Anti-glomerular basement membrane (anti-GBM) disease may coexist with antineutrophil cytoplasmic antibodies (ANCA), a clinicopathologic entity termed double-positive disease, in which diffuse alveolar hemorrhage (DAH) is common but histologic confirmation of pulmonary capillaritis remains difficult. A 93-year-old woman presented with rapidly progressive renal failure, and serologic evaluation revealed markedly elevated myeloperoxidase (MPO)-ANCA and anti-GBM antibody levels. Despite corticosteroid therapy, the patient died of progressive renal failure. At autopsy, the kidneys demonstrated diffuse crescentic glomerulonephritis with linear IgG deposition along the glomerular basement membrane, consistent with anti-GBM nephritis, together with multiple cortical scars composed of clusters of globally sclerotic glomeruli and interstitial fibrosis. The lungs exhibited DAH with focal neutrophilic infiltration along the alveolar septa, accompanied by focal destruction of capillary walls with subtle leukocytoclasia. Immunohistochemical analysis using epithelial marker (cytokeratin AE1/AE3), endothelial markers (CD31, CD34, and ERG), and platelet markers (CD61 and CD42b) highlighted alveolar epithelial injury, loss of endothelial continuity, and extravascular platelet deposition, respectively, in areas where capillaritis was suspected. Spatial concordance of these findings supported the presence of structural injury associated with pulmonary capillaritis. This case highlights the diagnostic importance of rigorous histopathologic assessment complemented by combined immunohistochemistry in patients with DAH.

Lung adenocarcinoma exhibits diverse pathological features, some of which for example, histological grading and tumor spread through air spaces (STAS), can be used for prognosis. Extra-alveolar invasion (EX) is a unique pathological factor focused on depth of invasion, and is defined as tumor infiltration into periarterial, peribronchiolar, or interlobular septal stroma. We recently suggested EX as a prognostic indicator, but its significance in sublobar resection remains unclear. We investigated the prognostic impact of EX in 116 patients with pT1mi-pT2a non-mucinous lung adenocarcinoma treated by segmentectomy or wedge resection. EX was evaluated using elastic fiber staining. Among 10 recurrent cases, 8 were EX-positive, and 2 were EX-negative and STAS-positive. The EX(-)/STAS(+) cases had local recurrence, while the EX(+) cases exhibited metastatic recurrence. The pT1abc EX(+) group showed significantly shorter recurrence-free survival (RFS) than the pT1abc EX(-) group, but no significant differences were observed when compared with the pT2 group. EX(+ )/STAS(-) and EX(+ )/STAS(+) groups showed significantly shorter RFS than EX(-)/STAS(-) group, but EX(-)/STAS(+) group did not. In conclusion, EX was associated with poor prognosis after sublobar resection of lung adenocarcinoma. Combined evaluation of EX and STAS may improve postoperative risk stratification.

IgG4-related ophthalmic disease (IgG4-ROD) is a common manifestation of IgG4-related disease, but its clinicopathologic features and the mechanisms underlying fibrosis remain incompletely understood. We retrospectively analyzed 100 cases of IgG4-ROD and investigated fibrosis using immunohistochemistry for IgG4, FOXP3, CD68, and CD163, together with RNA in situ hybridization for TGFB1. IgG4-ROD was histologically categorized into four patterns (Patterns 1-4) reflecting progressive fibrosis, demonstrating a spectrum from lymphoplasmacytic inflammation to marked fibrosis. Fibrosis was absent or minimal in 51% of cases, indicating that fibrosis is not essential for diagnosis, whereas obliterative phlebitis was rare. Compared with non-fibrotic cases, fibrotic cases showed significantly increased numbers of CD68-positive and CD163-positive macrophages (p < 0.001), predominantly localized at the interface between lymphoid and fibrotic areas. In contrast, fibrosis was not associated with IgG4-positive plasma cells, FOXP3-positive regulatory T cells, or EBV status. TGFB1 expression was significantly higher in lymphocyte-rich than fibrotic areas (p < 0.001), suggesting a closer association with lymphocyte-rich inflammation than established fibrosis. These findings demonstrate progressive histologic changes from Pattern 1 to Pattern 4 and suggest that accumulation of CD163-positive macrophages is associated with fibrosis in IgG4-ROD.

Pleural mesothelioma is an aggressive tumor with a poor prognosis and limited therapeutic options. While histological heterogeneity is common, true squamous differentiation is extremely rare and diagnostically challenging. We report an 81-year-old man with asbestos exposure who presented with recurrent right-sided pleural effusion. Thoracoscopic biopsies revealed two distinct tumor components. The first expressed mesothelial immunohistochemical markers, whereas the second showed a squamous profile. Both components demonstrated loss of BAP1 expression and homozygous deletion of CDKN2A, supporting a common mesothelial origin. Whole-exome sequencing revealed a tumor mutational burden above the average reported for mesothelioma (8-9 mutations per megabase), with 69 somatic mutations in total, 43 shared between the two components. In addition to canonical mesothelioma alterations (BAP1, CHEK2), the squamous component harbored 18 unique variants, including oncogenic alterations in NFE2L2 and PIK3CA. A novel SOX9 missense variant of uncertain significance (p. Thr243Pro) was also identified within the transactivation middle (TAM) domain; its functional consequence remains unknown. This case highlights a rare histological variant of pleural mesothelioma with partially squamous differentiation, characterized by a mutational burden above the mesothelioma average and squamous carcinoma-associated alterations. Integrated histology, immunohistochemistry, and genomic profiling are essential to avoid misdiagnosis and to improve understanding of tumor plasticity in mesothelioma.

Perivascular epithelioid cell tumors (PEComas) are mesenchymal neoplasms characterized by melanocytic and smooth muscle differentiation, commonly associated with TSC1/TSC2 alterations or TFE3 rearrangements. Herein, we report a rare case of urinary bladder PEComa with MITF overexpression and without identifiable TSC1/TSC2 mutations or TFE3 rearrangement. A 50-year-old man underwent partial cystectomy for a bladder mass. Histologically, the tumor was well circumscribed and composed of mixed spindle and epithelioid cells with cytologic atypia and low mitotic activity. Immunohistochemistry revealed diffuse expression of SMA, HMB45, and MITF, with negative staining for TFE3. Quantitative reverse-transcription PCR revealed increased MITF mRNA expression in PEComa tissue. Next-generation sequencing identified a truncating ATRX mutation but no alterations in TSC1/TSC2 or TFE3, and no MITF copy number gain. By Folpe criteria, the tumor was classified as malignant, whereas alternative models suggested a lower-risk category, highlighting the limitations of histologic risk stratification systems. The absence of canonical TSC/mTOR or TFE3 alterations, together with MITF overexpression, suggests a distinct molecular mechanism potentially involving dysregulation of the MiT/TFE pathway. This case expands the molecular spectrum of PEComa and supports the existence of a MITF-driven subset, highlighting the need for integrated histologic and molecular evaluation for prognostic assessment.

Glioblastoma, IDH-wildtype, is the most common and malignant adult-type diffuse glioma. It is characterized by extensive heterogeneity and poor prognosis. Recent single-cell RNA-sequencing studies have revealed distinct transcriptional programs that define cellular states such as astrocyte-like, mesenchymal-like, neural-progenitor-cell-like, and oligodendrocyte-progenitor-cell-like. However, their spatial context remains unclear because of the limitations of single-cell and spot-based spatial transcriptomics. In the present study, we applied a single-cell molecular classification of glioblastoma to a spatial transcriptomic analysis using the Xenium in situ platform. A custom gene panel comprising 366 genes, including 100 additional genes beyond the predesigned central nervous system panel, was designed to enable high-resolution, cell-level analysis. We constructed reduced modules derived from the Neftel et al. classification and validated their reproducibility across public single-cell datasets, achieving approximately 80% concordance with the original modules. In three cases of glioblastoma, IDH-wildtype, spatial transcriptomic profiling revealed regional variations in transcriptional states; cellular tumor regions, regions of pseudopalisading necrosis were significantly enriched in reduced neural-progenitor-like-cells subtype 1, and reduced mesenchymal-like-cells subtype 2, respectively. Our data indicate that the novel reduced module provides a framework for investigating single-cell molecular states within a tissue context in glioblastoma.

Tumors grow because the total number of cancer cells increases without limit. The prevailing explanation is that each proliferating cancer cell has acquired unlimited replicative capacity, yet this does not explain why the overwhelming majority of cancer cells fail to initiate tumors on transplantation, why the cancer stem cell fraction of serially passaged cultures remains nearly constant, or why the measured cell loss factor of human tumors is so high. This review proposes an alternative reading of these observations. Non-stem cancer cells may possess a replicative lifespan that is set not by telomere attrition but by the progressive accumulation of damaged proteins, protein aggregates and dysfunctional mitochondria, so that each lineage eventually terminates in growth arrest or death. Persistent expansion would then depend on th e rate at which non-stem cancer cells are converted back into stem cells. Work on transforming growth factor-β associated molecules, including TMEPAI/PMEPA1, MAFK-GPNMB and THG-1, is reviewed in this light, together with evidence from budding yeast and mammalian stem cells that damage is partitioned asymmetrically at division. Established observations, reasonable inferences and untested assumptions are distinguished explicitly, and experimental tests are proposed.

A man in his 50s developed persistent ventilator dependence, recurrent pneumonia, and lung abscess after right single-lung transplantation, and later died following complications from native lung pneumonectomy. Autopsy confirmed the diagnosis of tracheomalacia, allowing detailed histological examination that revealed extensive cartilage degeneration, disruption, and fibrosis of the trachea. Because histopathological studies of adult tracheomalacia are extremely rare, this case provides valuable insight into its pathology.

We herein review the pathological diagnosis of nodal T follicular helper (TFH) cell lymphoma, angioimmunoblastic type (nTFHL-AI), a mature T-cell neoplasm with a TFH phenotype that was previously designated as angioimmunoblastic T-cell lymphoma (AITL). The history of AITL from its discovery to the WHO 5th classification, which includes Japanese pathologists' and hematologists' contributions, is discussed. We then focus on what true neoplastic T cells of nTFHL-Al are in histopathology. Since the number of neoplastic cells is small in some cases of nTFHL-AI, the findings of recent single-cell analyses to immunohistochemistry of TFH markers are considered. We also discuss Epstein-Barr virus large B-cells in AITL, particularly diffuse large B-cell lymphoma and classic Hodgkin lymphoma (CHL), and propose how to make a differential diagnosis between nTFHL-Al and CHL.

Pleural mesothelioma (PM) often presents loss of BAP1 and MTAP genes. Digital PCR (dPCR) can accurately measure gene copy numbers (CNs). We hypothesized that cell-free DNA (cfDNA) extracted from the supernatant of pleural fluids (SPFs) could allow an efficient detection of these genetic events. BAP1/MTAP CN was measured with dPCR in SPFs and in solid tumor biopsies (STBs) of 37 PM patients. Results were compared with immunohistochemistry (IHC) carried out in STBs. IHC alone detected BAP1/MTAP-losses in 32/37 (86.5%). However, the sensitivity improved to 34/37 (91.9%) when low CN of BAP1/MTAP was included. Among 20 patients presenting SPFs, dPCR alone detected a low CN in 18 samples (sensitivity = 90%). Finally, this two-gene signature was applied on SPFs in a prospective series of six consecutive patients presenting an undefined diagnosis (negative for PM) and two showed reduced CN. After 7 months of follow-up, these subjects were further re-evaluated for the persistence/worsening of the symptoms and were diagnosed for PM. We highlighted dPCR's potential for detecting subtle BAP1/MTAP losses in SPFs of PM patients. This appears as a complementary approach for a more accurate diagnosis in specific contexts, when high-risk patients receive negative results after IHC analysis.