Invasion and Grading of Pulmonary Non-Mucinous Adenocarcinoma.

Lung adenocarcinoma staging and grading were recently updated to reflect the link between histologic growth patterns and outcomes. The lepidic growth pattern is regarded as "in-situ," whereas all other patterns are regarded as invasive, though with stratification. Solid, micropapillary, and complex glandular patterns are associated with worse prognosis than papillary and acinar patterns. These recent changes have improved prognostic stratification. However, multiple pitfalls exist in measuring invasive size and in classifying lung adenocarcinoma growth patterns. Awareness of these limitations and recommended practices will help the pathology community achieve consistent prognostic performance and potentially contribute to improved patient management.

The Grading System for Lung Adenocarcinoma: Brief Review of its Prognostic Performance and Future Directions.

Histologic grading of tumors is associated with prognosis in many organs. In the lung, the most recent grading system proposed by International association for the Study of Lung Cancer (IASLC) and adopted by the World Health Organization (WHO) incorporates the predominant histologic pattern, as well as the presence of high-grade architectural patterns (solid, micropapillary, and complex glandular pattern) in proportions >20% of the tumor surface. This system has shown improved prognostic ability when compared with the prior grading system based on the predominant pattern alone, across different patient populations. Interobserver agreement is moderate to excellent, depending on the study. IASLC/WHO grading system has been shown to correlate with molecular alterations and PD-L1 expression in tumor cells. Recent studies interrogating gene expression has shown correlation with tumor grade and molecular alterations in the tumor microenvironment that can further stratify risk of recurrence. The use of machine learning algorithms to grade nonmucinous adenocarcinoma under this system has shown accuracy comparable to that of expert pulmonary pathologists. Future directions include evaluation of tumor grade in the context of adjuvant and neoadjuvant therapies, as well as the development of better prognostic indicators for mucinous adenocarcinoma.

Adeno-to-squamous transition drives resistance to KRAS inhibition in LKB1 mutant lung cancer.

KRASG12C inhibitors (adagrasib and sotorasib) have shown clinical promise in targeting KRASG12C-mutated lung cancers; however, most patients eventually develop resistance. In lung patients with adenocarcinoma with KRASG12C and STK11/LKB1 co-mutations, we find an enrichment of the squamous cell carcinoma gene signature in pre-treatment biopsies correlates with a poor response to adagrasib. Studies of Lkb1-deficient KRASG12C and KrasG12D lung cancer mouse models and organoids treated with KRAS inhibitors reveal tumors invoke a lineage plasticity program, adeno-to-squamous transition (AST), that enables resistance to KRAS inhibition. Transcriptomic and epigenomic analyses reveal ΔNp63 drives AST and modulates response to KRAS inhibition. We identify an intermediate high-plastic cell state marked by expression of an AST plasticity signature and Krt6a. Notably, expression of the AST plasticity signature and KRT6A at baseline correlates with poor adagrasib responses. These data indicate the role of AST in KRAS inhibitor resistance and provide predictive biomarkers for KRAS-targeted therapies in lung cancer.

International reproducibility study of thymic epithelial tumors staging: pT stage is an issue. proposals for improvement. A RYTHMIC/ITMIG study.

INTRODUCTION: Pathologists are staging thymic epithelial tumors (TET) according to the 8th UICC/AJCC TNM system. Within the French RYTHMIC network, dedicated to TET, agreement on pathologic tumor stage (pT) among the pathology panelists was difficult. The aim of our study was to determine the interobserver reproducibility of pT at an international level, to explore the source of discrepancies and potential interventions to address these. METHODS: An international panel of pathologists was recruited through the International Thymic Malignancy Interest Group (ITMIG). The study focused on invasion of mediastinal pleura, pericardium, and lung. From a cohort of cases identified as challenging within the RYTHMIC network, we chose a series of test and validation cases (n = 5 and 10, respectively). RESULTS: Reproducibility of the pT stage was also challenging at an international level as none of the 15 cases was classified as the same pT stage by all ITMIG pathologists. The agreement rose from slight (κ = 0.13) to moderate (κ = 0.48) between test and validation series. Discussion among the expert pathologists pinpointed two major reasons underlying discrepancies: 1) Thymomas growing with their "capsule" and adhering to the pleurae, pericardium, or lung were often misinterpreted as invading these structures. 2) Recognition of the mediastinal pleura was identified as challenging. CONCLUSION: Our study underlines that the evaluation of the pT stage of TET is problematic and needs to be addressed in more detail in an upcoming TNM classification. The publication of histopathologic images of landmarks, including ancillary tests could improve reproducibility for future TNM classifications.

Regulatory T cells and IFN-γ-producing Th1 cells play a critical role in the pathogenesis of Sjögren's Syndrome.

Objectives: Sjögren's Disease (SjD) is an autoimmune disorder characterized by progressive dysfunction, inflammation and destruction of salivary and lacrimal glands, and by extraglandular manifestations. Its etiology and pathophysiology remain incompletely understood, though a role for autoreactive B cells has been considered key. Here, we investigated the role of effector and regulatory T cells in the pathogenesis of SjD. Methods: Histological analysis, RNA-sequencing and flow cytometry were conducted on glands, lungs, eyes and lymphoid tissues of mice with regulatory T cell-specific deletion of stromal interaction proteins (STIM) 1 and 2 ( Stim1/2 Foxp3 ), which play key roles in calcium signaling and T cell function. The pathogenicity of T cells from Stim1/2 Foxp3 mice was investigated through adoptively transfer into lymphopenic host mice. Additionally, single-cell transcriptomic analysis was performed on peripheral blood mononuclear cells (PBMCs) of patients with SjD and control subjects. Results: Stim1/2 Foxp3 mice develop a severe SjD-like disorder including salivary gland (SG) and lacrimal gland (LG) inflammation and dysfunction, autoantibodies and extraglandular symptoms. SG inflammation in Stim1/2 Foxp3 mice is characterized by T and B cell infiltration, and transcriptionally by a Th1 immune response that correlates strongly with the dysregulation observed in patients with SjD. Adoptive transfer of effector T cells from Stim1/2 Foxp3 mice demonstrates that the SjD-like disease is driven by interferon (IFN)-γ producing autoreactive CD4 + T cells independently of B cells and autoantiboodies. scRNA-seq analysis identifies increased Th1 responses and attenuated memory Treg function in PBMCs of patients with SjD. Conclusions: We report a more accurate mouse model of SjD while providing evidence for a critical role of Treg cells and IFN-γ producing Th1 cells in the pathogenesis of SjD, which may be effective targets for therapy.

Updates on lung adenocarcinoma: invasive size, grading and STAS.

Advancements in the classification of lung adenocarcinoma have resulted in significant changes in pathological reporting. The eighth edition of the tumour-node-metastasis (TNM) staging guidelines calls for the use of invasive size in staging in place of total tumour size. This shift improves prognostic stratification and requires a more nuanced approach to tumour measurements in challenging situations. Similarly, the adoption of new grading criteria based on the predominant and highest-grade pattern proposed by the International Association for the Study of Lung Cancer (IASLC) shows improved prognostication, and therefore clinical utility, relative to previous grading systems. Spread through airspaces (STAS) is a form of tumour invasion involving tumour cells spreading through the airspaces, which has been highly researched in recent years. This review discusses updates in pathological T staging, adenocarcinoma grading and STAS and illustrates the utility and limitations of current concepts in lung adenocarcinoma.

Inflammation in the tumor-adjacent lung as a predictor of clinical outcome in lung adenocarcinoma.

Approximately 30% of early-stage lung adenocarcinoma patients present with disease progression after successful surgical resection. Despite efforts of mapping the genetic landscape, there has been limited success in discovering predictive biomarkers of disease outcomes. Here we performed a systematic multi-omic assessment of 143 tumors and matched tumor-adjacent, histologically-normal lung tissue with long-term patient follow-up. Through histologic, mutational, and transcriptomic profiling of tumor and adjacent-normal tissue, we identified an inflammatory gene signature in tumor-adjacent tissue as the strongest clinical predictor of disease progression. Single-cell transcriptomic analysis demonstrated the progression-associated inflammatory signature was expressed in both immune and non-immune cells, and cell type-specific profiling in monocytes further improved outcome predictions. Additional analyses of tumor-adjacent transcriptomic data from The Cancer Genome Atlas validated the association of the inflammatory signature with worse outcomes across cancers. Collectively, our study suggests that molecular profiling of tumor-adjacent tissue can identify patients at high risk for disease progression.

KEAP1 mutation in lung adenocarcinoma promotes immune evasion and immunotherapy resistance.

Lung cancer treatment has benefited greatly through advancements in immunotherapies. However, immunotherapy often fails in patients with specific mutations like KEAP1, which are frequently found in lung adenocarcinoma. We established an antigenic lung cancer model and used it to explore how Keap1 mutations remodel the tumor immune microenvironment. Using single-cell technology and depletion studies, we demonstrate that Keap1-mutant tumors diminish dendritic cell and T cell responses driving immunotherapy resistance. This observation was corroborated in patient samples. CRISPR-Cas9-mediated gene targeting revealed that hyperactivation of the NRF2 antioxidant pathway is responsible for diminished immune responses in Keap1-mutant tumors. Importantly, we demonstrate that combining glutaminase inhibition with immune checkpoint blockade can reverse immunosuppression, making Keap1-mutant tumors susceptible to immunotherapy. Our study provides new insight into the role of KEAP1 mutations in immune evasion, paving the way for novel immune-based therapeutic strategies for KEAP1-mutant cancers.

Dataset for reporting of thymic epithelial tumours: recommendations from the International Collaboration on Cancer Reporting (ICCR).

AIMS: Thymic epithelial tumours (TET), including thymomas and thymic carcinomas and thymic neuroendocrine neoplasms, are malignant neoplasms that can be associated with morbidity and mortality. Recently, an updated version of the World Health Organization (WHO) Classification of Thoracic Tumours 5th Edition, 2021 has been released, which included various changes to the classification of these neoplasms. In addition, in 2017 the Union for International Cancer Control (UICC) / American Joint Committee on Cancer (AJCC) published the 8th Edition Staging Manual which, for the first time, includes a TNM staging that is applicable to thymomas, thymic carcinomas, and thymic neuroendocrine neoplasms. METHODS AND RESULTS: To standardize reporting of resected TET and thymic neuroendocrine neoplasms the accrediting bodies updated their reporting protocols. The International Collaboration on Cancer Reporting (ICCR), which represents a collaboration between various National Associations of Pathology, updated its 2017 histopathology reporting guide on TET and thymic neuroendocrine neoplasms accordingly. This report will highlight important changes in the reporting of TET and thymic neuroendocrine neoplasms based on the 2021 WHO, emphasize the 2017 TNM staging, and also comment on the rigour and various uncertainties for the pathologist when trying to follow that staging. CONCLUSION: The ICCR dataset provides a comprehensive, standardized template for reporting of resected TET and thymic neuroendocrine neoplasms.

Pathomic Features Reveal Immune and Molecular Evolution From Lung Preneoplasia to Invasive Adenocarcinoma.

Recent statistics on lung cancer, including the steady decline of advanced diseases and the dramatically increasing detection of early-stage diseases and indeterminate pulmonary nodules, mark the significance of a comprehensive understanding of early lung carcinogenesis. Lung adenocarcinoma (ADC) is the most common histologic subtype of lung cancer, and atypical adenomatous hyperplasia is the only recognized preneoplasia to ADC, which may progress to adenocarcinoma in situ (AIS) and minimally invasive adenocarcinoma (MIA) and eventually to invasive ADC. Although molecular evolution during early lung carcinogenesis has been explored in recent years, the progress has been significantly hindered, largely due to insufficient materials from ADC precursors. Here, we employed state-of-the-art deep learning and artificial intelligence techniques to robustly segment and recognize cells on routinely used hematoxylin and eosin histopathology images and extracted 9 biology-relevant pathomic features to decode lung preneoplasia evolution. We analyzed 3 distinct cohorts (Japan, China, and United States) covering 98 patients, 162 slides, and 669 regions of interest, including 143 normal, 129 atypical adenomatous hyperplasia, 94 AIS, 98 MIA, and 205 ADC. Extracted pathomic features revealed progressive increase of atypical epithelial cells and progressive decrease of lymphocytic cells from normal to AAH, AIS, MIA, and ADC, consistent with the results from tissue-consuming and expensive molecular/immune profiling. Furthermore, pathomics analysis manifested progressively increasing cellular intratumor heterogeneity along with the evolution from normal lung to invasive ADC. These findings demonstrated the feasibility and substantial potential of pathomics in studying lung cancer carcinogenesis directly from the low-cost routine hematoxylin and eosin staining.

Automated and robust extraction of genomic DNA from various leftover blood samples.

With the development of genomic technologies, the isolation of genomic DNA (gDNA) from clinical samples is increasingly required for clinical diagnostics and research studies. In this study, we explored the potential of utilizing various leftover blood samples obtained from routine clinical tests as a viable source of gDNA. Using an automated method with optimized pre-treatments, we obtained gDNA from seven types of clinical leftover blood, with average yields of gDNA ranging from 3.11 ± 0.45 to 22.45 ± 4.83 µg. Additionally, we investigated the impact of storage conditions on gDNA recovery, resulting in yields of 8.62-68.08 µg when extracting gDNA from EDTA leftover blood samples stored at 4 °C for up to 13 weeks or -80 °C for up to 78 weeks. Furthermore, we successfully obtained sequenceable gDNA from both Serum Separator Tube and EDTA Tube using a 96-well format extraction, with yields ranging from 0.61 to 71.29 µg and 3.94-215.98 µg, respectively. Our findings demonstrate the feasibility of using automated high-throughput platforms for gDNA extraction from various clinical leftover blood samples with the proper pre-treatments.

Deep learning integrates histopathology and proteogenomics at a pan-cancer level.

We introduce a pioneering approach that integrates pathology imaging with transcriptomics and proteomics to identify predictive histology features associated with critical clinical outcomes in cancer. We utilize 2,755 H&E-stained histopathological slides from 657 patients across 6 cancer types from CPTAC. Our models effectively recapitulate distinctions readily made by human pathologists: tumor vs. normal (AUROC = 0.995) and tissue-of-origin (AUROC = 0.979). We further investigate predictive power on tasks not normally performed from H&E alone, including TP53 prediction and pathologic stage. Importantly, we describe predictive morphologies not previously utilized in a clinical setting. The incorporation of transcriptomics and proteomics identifies pathway-level signatures and cellular processes driving predictive histology features. Model generalizability and interpretability is confirmed using TCGA. We propose a classification system for these tasks, and suggest potential clinical applications for this integrated human and machine learning approach. A publicly available web-based platform implements these models.

Tumor-intrinsic LKB1-LIF signaling axis establishes a myeloid niche to promote immune evasion and tumor growth.

Tumor mutations can influence the surrounding microenvironment leading to suppression of anti-tumor immune responses and thereby contributing to tumor progression and failure of cancer therapies. Here we use genetically engineered lung cancer mouse models and patient samples to dissect how LKB1 mutations accelerate tumor growth by reshaping the immune microenvironment. Comprehensive immune profiling of LKB1 -mutant vs wildtype tumors revealed dramatic changes in myeloid cells, specifically enrichment of Arg1 + interstitial macrophages and SiglecF Hi neutrophils. We discovered a novel mechanism whereby autocrine LIF signaling in Lkb1 -mutant tumors drives tumorigenesis by reprogramming myeloid cells in the immune microenvironment. Inhibiting LIF signaling in Lkb1 -mutant tumors, via gene targeting or with a neutralizing antibody, resulted in a striking reduction in Arg1 + interstitial macrophages and SiglecF Hi neutrophils, expansion of antigen specific T cells, and inhibition of tumor progression. Thus, targeting LIF signaling provides a new therapeutic approach to reverse the immunosuppressive microenvironment of LKB1 -mutant tumors.

Bifidobacterium animalis subspecies lactis HN019 can reduce the sequelae of experimental periodontitis in rats modulating intestinal parameters, expression of lipogenic genes, and levels of hepatic steatosis.

OBJECTIVE: To determine whether Bifidobacterium animalis subspecies lactis HN019 (B. lactis HN019) can reduce the sequelae of experimental periodontitis (EP) in rats modulating systemic parameters. BACKGROUND: This study evaluated the effects of probiotic therapy (PROB) in the prevention of local and systemic damage resulting from EP. METHODS: Forty-eight rats were allocated into four groups: C (control), PROB, EP, and EP-PROB. PROB (1 × 1010 CFU/mL) administration lasted 8 weeks and PE was induced on the 7th week by placing ligature on the animals' lower first molars. All animals were euthanized in the 9th week of the experiment. Biomolecular analyses, RT-PCR, and histomorphometric analyses were performed. The data obtained were analyzed statistically (ANOVA, Tukey, p < .05). RESULTS: The EP group had higher dyslipidemia when compared to the C group, as well as higher levels of insulin resistance, proteinuria levels, percentages of systolic blood pressure, percentage of fatty hepatocytes in the liver, and expression of adipokines was up-regulated (LEPR, NAMPT, and FABP4). All these parameters (except insulin resistance, systolic blood pressure, LEPR and FABP4 gene expression) were reduced in the EP-PROB group when compared to the EP group. The EP group had lower villus height and crypt depth, as well as a greater reduction in Bacteroidetes and a greater increase in Firmicutes when compared to the EP-PROB group. Greater alveolar bone loss was observed in the EP group when compared to the EP-PROB group. CONCLUSION: Bifidobacterium lactis HN019 can reduce the sequelae of EP in rats modulating intestinal parameters, attenuating expression of lipogenic genes and hepatic steatosis.

Correlation of Programmed Death-Ligand 1 Expression With Lung Adenocarcinoma Histologic and Molecular Subgroups in Primary and Metastatic Sites.

Programmed death-ligand 1 (PD-L1) expression in terms of the tumor proportion score (TPS) is the main predictive biomarker approved for immunotherapy against lung nonsmall cell carcinoma. Although some studies have explored the associations between histology and PD-L1 expression in pulmonary adenocarcinoma, they have been limited in sample size and/or extent of examined histologic variables, which may have resulted in conflicting information. In this observational retrospective study, we identified primary and metastatic lung adenocarcinoma cases in the span of 5 years and tabulated the detailed histopathologic features, including pathological stage, tumor growth pattern, tumor grade, lymphovascular and pleural invasion, molecular alterations, and the associated PD-L1 expression for each case. Statistical analyses were performed to detect associations between PD-L1 and these features. Among 1658 cases, 643 were primary tumor resections, 751 were primary tumor biopsies, and 264 were metastatic site biopsies or resections. Higher TPS significantly correlated with high-grade growth patterns, grade 3 tumors, higher T and N stage, presence of lymphovascular invasion, and presence of MET and TP53 alterations, whereas lower TPS correlated with lower-grade tumors and presence of EGFR alterations. There was no difference in PD-L1 expression in matched primary and metastases, although higher TPS was observed in metastatic tumors due to the presence of high-grade patterns in these specimens. TPS showed a strong association with a histologic pattern. Higher-grade tumors had higher TPS, which is also associated with more aggressive histologic features. Tumor grade should be kept in mind when selecting cases and blocks for PD-L1 testing.

Histopathology of the Mitral Valve Residual Leaflet in Obstructive Hypertrophic Cardiomyopathy.

BACKGROUND: Mitral valve (MV) elongation is a primary hypertrophic cardiomyopathy (HCM) phenotype and contributes to obstruction. The residual MV leaflet that protrudes past the coaptation point is especially susceptible to flow-drag and systolic anterior motion. Histopathological features of MVs in obstructive hypertrophic cardiomyopathy (OHCM), and of residual leaflets specifically, are unknown. OBJECTIVES: The purpose of this study was to characterize gross, structural, and cellular histopathologic features of MV residual leaflets in OHCM. On a cellular-level, we assessed for developmental dysregulation of epicardium-derived cell (EPDC) differentiation, adaptive endocardial-to-mesenchymal transition and valvular interstitial cell proliferation, and genetically-driven persistence of cardiomyocytes in the valve. METHODS: Structural and immunohistochemical staining were performed on 22 residual leaflets excised as ancillary procedures during myectomy, and compared with 11 control leaflets from deceased patients with normal hearts. Structural components were assessed with hematoxylin and eosin, trichrome, and elastic stains. We stained for EPDCs, EPDC paracrine signaling, valvular interstitial cells, endocardial-to-mesenchymal transition, and cardiomyocytes. RESULTS: The residual leaflet was always at A2 segment and attached by slack, elongated and curlicued, myxoid chords. MV residual leaflets in OHCM were structurally disorganized, with expanded spongiosa and increased, fragmented elastic fibers compared with control leading edges. The internal collagenous fibrosa was attenuated and there was collagenous tissue overlying valve surfaces in HCM, with an overall trend toward decreased leaflet thickness (1.09 vs 1.47 mm, P = 0.08). No markers of primary cellular processes were identified. CONCLUSIONS: MV residual leaflets in HCM were characterized by histologic findings that were likely secondary to chronic hemodynamic stress and may further increase susceptibility to systolic anterior motion.

Case report: Primary adenocarcinoma NOS of the thymus and cytological features.

Aspirates of mediastinal neoplasms pose a unique diagnostic challenge due to the overlapping histologic characteristics of mediastinal lesions and the morphologic similarities between mediastinal neoplasms and those originating at other sites. Presented here is the first reported description of the cytomorphologic features of adenocarcinoma NOS of the thymus in aspirate and pleural effusion specimens. The morphologic similarities between thymic and metastatic adenocarcinomas and variable immunohistochemical staining patterns of thymic epithelial neoplasms underscore the importance of pathology-radiology correlation and the careful consideration of the clinical context in the interpretation of cytology specimens.

Effects of probiotics in rats with experimental metabolic syndrome and periodontitis: An investigation of the intestine-adipose tissue axis.

BACKGROUND: This study evaluated the systemic (intestine and adipose tissue) and local (periodontal tissues) impact of probiotic therapy in rats with metabolic syndrome (MS) associated or not with periodontitis (PE). METHODS: Forty-eight rats received a high-fat diet for induction of MS for 16 weeks. They were subdivided into groups with (+) and without (-) PE, receiving (*) or not (**) receiving probiotics (PROB): MS (-**), MSP (-*), MSPE (+**), and MSPEP (+*). PROB administration (Bifidobacterium animalis subsp. lactis HN019) started on the 8th week of the study and PE was induced on the 14th week by placing ligature on the animals' lower first molars. Euthanasia occurred in the 16th week. Biomolecular, immunoenzymatic assays, and histomorphometric analyses were performed. The data obtained were statistically analyzed (ANOVA, Tukey, p < 0.05). RESULTS: The MSPEP group exhibited reduced alveolar bone loss when compared with the MSPE group, as well as lower levels of hepatic steatosis and proteinuria (p < 0.05). In the intestinal environment, the MSPE group exhibited significantly lower villus height and crypt depth, as well as a greater increase in Bacillota when compared with the MSPEP group (p < 0.05). The MSPEP group showed lower adipokine gene expression (LEPR, NAMPT, and FABP4) in adipose tissue than the MSPE group (p < 0.05). CONCLUSION: The probiotic B. lactis HN019 reduced the severity of experimental periodontitis and modulated the expression of lipogenic genes and intestinal morphological and microbiological parameters in rats with MS.

Extensive fibrosis in mediastinal seminoma is a diagnostic pitfall in small biopsies: two case reports.

Background: In mediastinal biopsies that show fibrosis, the differential diagnosis includes fibrosing mediastinitis, immunoglobulin G subclass 4-related disease, Hodgkin lymphoma, as well as reactive fibrotic and inflammatory changes adjacent to other processes including neoplasms. Cases Description: We report two cases of incidentally detected mediastinal seminoma that contained extensive areas of paucicellular fibrosis, which precluded accurate preoperative biopsy diagnosis. The fibrosis consisted of mildly inflamed, densely scarred tissue with thin dilated vessels, and was present to a significant extent that is suggestive of spontaneous regression. These features are not currently described in the World Health Organization Classification of Thoracic Tumors. In both patients, needle and open biopsies sampled only the fibrotic areas of the tumors, and the final diagnosis was not achieved until surgical excision was performed. After surgery, both patients received chemotherapy, and were alive without evidence of disease at 3.4 years and 1 year post-operatively, respectively. Tumor fibrosis composed approximately 95% and 50% of each patient's tumor, respectively. In one of the patients, correlation of the needle biopsy position with the positron emission tomography (PET) scan revealed that the biopsy needle had sampled a non-metabolically active portion of the tumor. Conclusions: While pathologic spontaneous regression is well-described in gonadal germ cell tumors, it is not well-reported in extragonadal locations. Prospective knowledge of this diagnostic pitfall and targeting PET-avid regions of the tumor may increase the diagnostic yield and help to avoid non-indicated surgical interventions.