Long-term prospective outcome data using EndoPredict as risk stratification and chemotherapy decision biomarker in hormone receptor-positive, HER2-negative early breast cancer.

PURPOSE: To report the prospective long-term outcome data of patients whose chemotherapy decision was guided by the EndoPredict test. METHODS: Patients with hormone receptor-positive HER2-negative early breast cancer with 0-3 positive lymph nodes were enrolled. The EndoPredict test was carried out on all tumor samples. Treatment compliance, local recurrence, distant metastases, and survival were evaluated. Associations of EPclin risk stratification with 5-year disease-free survival and distant metastasis-free survival were evaluated by time-to-event analysis. RESULTS: 368 consecutive patients were included in the analysis. Median follow-up was 8.2 years. EndoPredict allocated 238 (65%) in the low-risk and 130 (35%) patients in the high-risk group. Risk for disease recurrence or death in EPclin high-risk patients was twofold higher than in EPclin low-risk patients (hazard ratio [HR] 2.08; 95% CI 1.26-3.44; p = 0.004). EPclin low-risk patients had a 5-year disease-free survival of 95.3% (95% CI 92.6-98.0%). EPclin high-risk patients were at higher risk of developing distant metastases or death (HR 2.21; 95% CI 1.27-3.88; p = 0.005). EPclin high-risk patients who underwent chemotherapy had a 5-year DFS of 89.1% (95% CI 82.7-96.1%) in contrast to high-risk patients without chemotherapy (68.9%; 95% CI 56.2-84.5%; HR 0.46; 95% CI 0.23-0.95; p = 0.036). EPclin high-risk patients were at higher risk of experiencing distant metastases or death than EPclin low-risk patients regardless of menopausal status (premenopausal: HR 3.55; 95% CI 1.17-12.32; p = 0.025; postmenopausal: HR 1.92; 95% CI 0.99-3.7; p = 0.054). CONCLUSION: EndoPredict can guide decisions on adjuvant chemotherapy in early luminal breast cancer. EndoPredict risk stratification is also applicable in premenopausal women.

Chondrosarcoma evaluation using hematein-based x-ray staining and high-resolution 3D micro-CT: a feasibility study.

BACKGROUND: Chondrosarcomas are rare malignant bone tumors diagnosed by analyzing radiological images and histology of tissue biopsies and evaluating features such as matrix calcification, cortical destruction, trabecular penetration, and tumor cell entrapment. METHODS: We retrospectively analyzed 16 cartilaginous tumor tissue samples from three patients (51-, 54-, and 70-year-old) diagnosed with a dedifferentiated chondrosarcoma at the femur, a moderately differentiated chondrosarcoma in the pelvis, and a predominantly moderately differentiated chondrosarcoma at the scapula, respectively. We combined a hematein-based x-ray staining with high-resolution three-dimensional (3D) microscopic x-ray computed tomography (micro-CT) for nondestructive 3D tumor assessment and tumor margin evaluation. RESULTS: We detected trabecular entrapment on 3D micro-CT images and followed bone destruction throughout the volume. In addition to staining cell nuclei, hematein-based staining also improved the visualization of the tumor matrix, allowing for the distinction between the tumor and the bone marrow cavity. The hematein-based staining did not interfere with further conventional histology. There was a 5.97 ± 7.17% difference between the relative tumor area measured using micro-CT and histopathology (p = 0.806) (Pearson correlation coefficient r = 0.92, p = 0.009). Signal intensity in the tumor matrix (4.85 ± 2.94) was significantly higher in the stained samples compared to the unstained counterparts (1.92 ± 0.11, p = 0.002). CONCLUSIONS: Using nondestructive 3D micro-CT, the simultaneous visualization of radiological and histopathological features is feasible. RELEVANCE STATEMENT: 3D micro-CT data supports modern radiological and histopathological investigations of human bone tumor specimens. It has the potential for being an integrative part of clinical preoperative diagnostics. KEY POINTS: • Matrix calcifications are a relevant diagnostic feature of bone tumors. • Micro-CT detects all clinically diagnostic relevant features of x-ray-stained chondrosarcoma. • Micro-CT has the potential to be an integrative part of clinical diagnostics.

[Comparative pathology in oncology-Best practice]. / Vergleichende Pathologie in der onkologischen Forschung.

Comparative experimental pathology is a research field at the interface of human and veterinary medicine. It is focused on the comparative study of similarities and differences between spontaneous and experimentally induced diseases in animals (animal models) compared to human diseases. The use of animal models for studying human diseases is an essential component of biomedical research. Interdisciplinary teams with species-specific expertise should collaborate wherever possible and maintain close communication. Mutual openness, cooperation, and willingness to learn form the basis for a fruitful collaboration. Research projects jointly led by or involving both animal and human pathologists make a significant contribution to high-quality biomedical research. Such approaches are promising not only in oncological research, as outlined in this article, but also in other research areas where animal models are regularly used, such as infectiology, neurology, and developmental biology.

[Personalized medicine in oncology]. / Personalisierte Medizin in der Onkologie.

Due to the considerable technological progress in molecular and genetic diagnostics as well as increasing insights into the molecular pathogenesis of diseases, there has been a fundamental paradigm shift in the past two decades from a "one-size-fits-all approach" to personalized, molecularly informed treatment strategies. Personalized medicine or precision medicine focuses on the genetic, physiological, molecular, and biochemical differences between individuals and considers their effects on the development, prevention, and treatment of diseases. As a pioneer of personalized medicine, the field of oncology is particularly noteworthy, where personalized diagnostics and treatment have led to lasting change in the treatment of cancer patients in recent years. In this article, the significant change towards personalized treatment concepts, especially in the field of personalized oncology, will be discussed and examined in more detail.

[Artificial intelligence for pathology-how, where, and why?] / Künstliche Intelligenz in der Pathologie ­ wie, wo und warum?

Artificial intelligence promises many innovations and simplifications in pathology, but also raises just as many questions and uncertainties. In this article, we provide a brief overview of the current status, the goals already achieved by existing algorithms, and the remaining challenges.

Endothelial cells drive organ fibrosis in mice by inducing expression of the transcription factor SOX9.

Fibrosis is a hallmark of chronic disease. Although fibroblasts are involved, it is unclear to what extent endothelial cells also might contribute. We detected increased expression of the transcription factor Sox9 in endothelial cells in several different mouse fibrosis models. These models included systolic heart failure induced by pressure overload, diastolic heart failure induced by high-fat diet and nitric oxide synthase inhibition, pulmonary fibrosis induced by bleomycin treatment, and liver fibrosis due to a choline-deficient diet. We also observed up-regulation of endothelial SOX9 in cardiac tissue from patients with heart failure. To test whether SOX9 induction was sufficient to cause disease, we generated mice with endothelial cell-specific overexpression of Sox9, which promoted fibrosis in multiple organs and resulted in signs of heart failure. Endothelial Sox9 deletion prevented fibrosis and organ dysfunction in the two mouse models of heart failure as well as in the lung and liver fibrosis mouse models. Bulk and single-cell RNA sequencing of mouse endothelial cells across multiple vascular beds revealed that SOX9 induced extracellular matrix, growth factor, and inflammatory gene expression, leading to matrix deposition by endothelial cells. Moreover, mouse endothelial cells activated neighboring fibroblasts that then migrated and deposited matrix in response to SOX9, a process partly mediated by the secreted growth factor CCN2, a direct SOX9 target; endothelial cell-specific Sox9 deletion reversed these changes. These findings suggest a role for endothelial SOX9 as a fibrosis-promoting factor in different mouse organs during disease and imply that endothelial cells are an important regulator of fibrosis.

Immunologic landscape of human hepatic hemangiomas and epithelioid hemangioendotheliomas.

BACKGROUND: The missing requirement for resection for the majority of hepatic hemangiomas (HH) and tissue scarcity for rare diseases such as hepatic epithelioid hemangioendotheliomas (HEHE) complicate the characterization of the spatial immunovascular niche of these benign and malignant vascular neoplastic diseases. METHODS: Two tissue cohorts containing 98 HHs and 13 HEHEs were used to study entity-specific and disease stage-specific endothelial cell (EC) phenotype and immune cell abundance. Using semiquantitative assessment, annotation-based cell classifiers, digital cell detection on whole slides, and tissue microarrays, we quantified 23 immunologic and vascular niche-associated markers and correlated this with clinicopathologic data. RESULTS: Both HH and HEHE ECs were characterized by a CD31high, CD34high, FVIII-related antigenhigh expression phenotype with entity-specific expression differences of sinusoidal EC markers Stabilin1, Stabilin2, CD32, and Lymphatic Vessel Endothelial Hyaluronan Receptor 1 (LYVE-1). Cell detection identified an HH margin-prevailing immunologic response dominated by Myeloperoxidase+ (MPO+) macrophages, CD3+ and CD8+ T cell subsets, and B cells (CD20+, CD79A+). In HEHE, increased CD68+ and CD20+ cell demarcation of lesion margins was observed, while CD3+ and CD8+ T cells were equally detectable both marginally and intralesionally. Stage-specific pairwise correlation analysis of HH and HEHE revealed disease entity-specific immunologic infiltration patterns as seen by high CD117+ cell numbers in HH, while HEHE samples showed increased CD3+ T cell infiltration. CONCLUSIONS: ECs in HH and HEHE share a continuous EC expression phenotype, while the expression of sinusoidal EC markers is more highly retained in HEHE. These phenotypic differences are associated with a unique and disease-specific immunovascular landscape.

[Digital transformation of a routine histopathology lab : Dos and don'ts!] / Digitalisierung der histopathologischen Routinediagnostik : Dos and Don'ts!

The implementation of digital histopathology in the laboratory marks a crucial milestone in the overall digital transformation of pathology. This shift offers a range of new possibilities, including access to extensive datasets for AI-assisted analyses, the flexibility of remote work and home office arrangements for specialists, and the expedited and simplified sharing of images and data for research, conferences, and tumor boards. However, the transition to a fully digital workflow involves significant technological and personnel-related efforts. It necessitates careful and adaptable change management to minimize disruptions, particularly in the personnel domain, and to prevent the loss of valuable potential from employees who may be resistant to change. This article consolidates our institute's experiences, highlighting technical and personnel-related challenges encountered during the transition to digital pathology. It also presents a comprehensive overview of potential difficulties at various interfaces when converting routine operations to a digital workflow.

CT Attenuation of Hepatic Pancreatic Cancer Metastases Correlates with Prognostically Detrimental Metastatic Necrosis.

Percutaneous CT-guided biopsy is a frequently performed procedure for the confirmation and molecular workup of hepatic metastases of pancreatic ductal adenocarcinoma (PDAC). Tumor necrosis of primary PDAC has shown a negative prognostic impact in recent studies. This study aims to examine predictability in CT scans and the prognostic impact of necrosis in hepatic metastases of PDAC. In this tertiary-center retrospective cohort study, we included 36 patients with hepatic metastases of PDAC who underwent CT-guided hepatic biopsies. Normalized attenuation of the biopsied metastasis was determined in venous phase contrast-enhanced planning scans obtained prior to biopsy by automatic, threshold-based 3D segmentation and manual, blinded 2D segmentation. A board-certified pathologist specialized in hepatic pathology histologically quantified the tumor necrosis and cellularity of the biopsy cylinders. We found a significant inverse-linear correlation between normalized attenuation and the fraction of necrosis (Pearson's r = 0.51, p < 0.001 for automatic 3D segmentation or Pearson's r = 0.52, p < 0.001 for manual 2D segmentation), whereas no correlation was found with tumor cellularity. Additionally, we discovered that patients with a fraction of necrosis ≥ 20% in metastases had a significantly shorter overall survival (p < 0.035). In summary, tumor necrosis of PDAC metastases can be estimated from contrast-enhanced CT scans, which could help to improve biopsy sample pattern planning. In addition, liver metastatic necrosis may serve as a prognostic biomarker in PDAC.

A mitochondria-regulated p53-CCF circuit integrates genome integrity with inflammation.

Genomic instability and inflammation are distinct hallmarks of aging, but the connection between them is poorly understood. Understanding their interrelationship will help unravel new mechanisms and therapeutic targets of aging and age-associated diseases. Here we report a novel mechanism directly linking genomic instability and inflammation in senescent cells, through a mitochondria-regulated molecular circuit that connects the p53 tumor suppressor and cytoplasmic chromatin fragments (CCF), a driver of inflammation through the cGAS-STING pathway. Activation or inactivation of p53 by genetic and pharmacologic approaches showed that p53 suppresses CCF accumulation and the downstream inflammatory senescence-associated secretory phenotype (SASP), independent of its effects on cell cycle arrest. p53 activation suppressed CCF formation by promoting DNA repair, reflected in maintenance of genomic integrity, particularly in subtelomeric regions, as shown by single cell genome resequencing. Activation of p53 by pharmacological inhibition of MDM2 in old mice decreased features of SASP in liver, indicating a senomorphic role in vivo . Remarkably, mitochondria in senescent cells suppressed p53 activity by promoting CCF formation and thereby restricting ATM-dependent nuclear DNA damage signaling. These data provide evidence for a mitochondria-regulated p53-CCF circuit in senescent cells that controls DNA repair, genome integrity and inflammatory SASP, and is a potential target for senomorphic healthy aging interventions.

Hepatocellular Brg1 promotes CCl4-induced liver inflammation, ECM accumulation and fibrosis in mice.

INTRODUCTION: Hepatic fibrosis is a progressive pathological process involving the exhaustion of hepatocellular regenerative capacity and ultimately leading to the development of cirrhosis and even hepatocellular carcinoma. Brg1, the core subunit of the SWI/SNF chromatin-remodeling complex, was recently identified as important for liver regeneration. This study investigates the role of Brg1 in hepatic fibrosis development. METHODS: Hepatocyte-specific Brg1 knockout mice were generated and injected with carbon tetrachloride (CCl4) for 4, 6, 8, and 12 weeks to induce liver fibrosis. Afterwards, liver fibrosis and liver damage were assessed. RESULTS: Brg1 expression was significantly increased in the fibrotic liver tissue of wild-type mice, as compared to that of untreated wild-type mice. The livers of the Brg1 knockout animals showed reduced liver inflammation, extracellular matrix accumulation, and liver fibrosis. TNF-α and NF-κB-mediated inflammatory response was reduced in Brg1 knockout animals. CONCLUSION: Brg1 promotes the progression of liver fibrosis in mice and may therefore be used as a potential therapeutic target for treating patients with liver fibrosis due to chronic injury.

Histomorphometric Analysis of 38 Giant Cell Tumors of Bone after Recurrence as Compared to Changes Following Denosumab Treatment.

Giant cell tumor of bone (GCTB) is an osteolytic tumor driven by an H3F3A-mutated mononuclear cell with the accumulation of osteoclastic giant cells. We analyzed tissue from 13 patients with recurrence and 25 patients with denosumab therapy, including two cases of malignant transformation. We found a decrease in the total number of cells (p = 0.03), but not in the individual cell populations when comparing primary and recurrence. The patients treated with denosumab showed induction of osteoid formation increasing during therapy. The total number of cells was reduced (p < 0.0001) and the number of H3F3A-mutated tumor cells decreased (p = 0.0001), while the H3F3A wild-type population remained stable. The KI-67 proliferation rate dropped from 10% to 1% and Runx2- and SATB2-positive cells were reduced. The two cases of malignant transformation revealed a loss of the H3F3A-mutated cells, while the KI-67 rate increased. Changes in RUNX2 and SATB2 expression were higher in one sarcoma, while in the other RUNX2 was decreased and SATB2-positive cells were completely lost. We conclude that denosumab has a strong impact on the morphology of GCTB. KI-67, RUNX2 and SATB2 expression differed depending on the benign or malignant course of the tumor under denosumab therapy.

Endothelial Notch1 signaling in white adipose tissue promotes cancer cachexia.

Cachexia is a major cause of morbidity and mortality in individuals with cancer and is characterized by weight loss due to adipose and muscle tissue wasting. Hallmarks of white adipose tissue (WAT) remodeling, which often precedes weight loss, are impaired lipid storage, inflammation and eventually fibrosis. Tissue wasting occurs in response to tumor-secreted factors. Considering that the continuous endothelium in WAT is the first line of contact with circulating factors, we postulated whether the endothelium itself may orchestrate tissue remodeling. Here, we show using human and mouse cancer models that during precachexia, tumors overactivate Notch1 signaling in distant WAT endothelium. Sustained endothelial Notch1 signaling induces a WAT wasting phenotype in male mice through excessive retinoic acid production. Pharmacological blockade of retinoic acid signaling was sufficient to inhibit WAT wasting in a mouse cancer cachexia model. This demonstrates that cancer manipulates the endothelium at distant sites to mediate WAT wasting by altering angiocrine signals.

Fulminant Liver Failure after Treatment with a Checkpoint Inhibitor for Gastric Cancer: A Case Report and Review of the Literature.

Nivolumab is a promising monoclonal antibody inhibitor of programmed death-1, a protein on the surface of T-cells. As such, it is approved for use in patients with multiple advanced malignancies and can significantly elongate progression-free survival. However, monoclonal antibody inhibitors can lead to adverse hepatic reactions, which in rare cases result in further hepatic damage. Herein, we present a case of a patient with locally advanced gastric carcinoma treated with fluorouracil, oxaliplatin, docetaxel and the checkpoint inhibitor nivolumab. Five months after her first dosage of nivolumab and without a preexisting liver disease, she presented with transaminitis. During the course of her stay, the patient developed status epilepticus, which required mechanical ventilation followed by fulminant hepatic failure. A subsequent liver biopsy revealed severe liver damage with extensive confluent parenchymal necrosis corresponding to checkpoint-inhibitor-induced hepatitis. Alternative reasons for this hepatic failure were ruled out. Despite aggressive therapeutic interventions including corticosteroids and plasma exchange, the patient died due to liver failure. Although hepatic failure is rarely seen in patients with checkpoint inhibitor therapy, it requires early awareness and rapid intervention.

Liver sinusoidal endothelial cells orchestrate NK cell recruitment and activation in acute inflammatory liver injury.

Liver sinusoidal endothelial cells (LSECs) rapidly clear lipopolysaccharide (LPS) from the bloodstream and establish intimate contact with immune cells. However, their role in regulating liver inflammation remains poorly understood. We show that LSECs modify their chemokine expression profile driven by LPS or interferon-γ (IFN-γ), resulting in the production of the myeloid- or lymphoid-attracting chemokines CCL2 and CXCL10, respectively, which accumulate in the serum of LPS-challenged animals. Natural killer (NK) cell exposure to LSECs in vitro primes NK cells for higher production of IFN-γ in response to interleukin-12 (IL-12) and IL-18. In livers of LPS-injected mice, NK cells are the major producers of this cytokine. In turn, LSECs require exposure to IFN-γ for CXCL10 expression, and endothelial-specific Cxcl10 gene deletion curtails NK cell accumulation in the inflamed livers. Thus, LSECs respond to both LPS and immune-derived signals and fuel a positive feedback loop of immune cell attraction and activation in the inflamed liver tissue.

Semaphorin 3C exacerbates liver fibrosis.

BACKGROUND AND AIMS: Chronic liver disease is a growing epidemic, leading to fibrosis and cirrhosis. TGF-ß is the pivotal profibrogenic cytokine that activates HSC, yet other molecules can modulate TGF-ß signaling during liver fibrosis. Expression of the axon guidance molecules semaphorins (SEMAs), which signal through plexins and neuropilins (NRPs), have been associated with liver fibrosis in HBV-induced chronic hepatitis. This study aims at determining their function in the regulation of HSCs. APPROACH AND RESULTS: We analyzed publicly available patient databases and liver biopsies. We used transgenic mice, in which genes are deleted only in activated HSCs to perform ex vivo analysis and animal models. SEMA3C is the most enriched member of the semaphorin family in liver samples from patients with cirrhosis. Higher expression of SEMA3C in patients with NASH, alcoholic hepatitis, or HBV-induced hepatitis discriminates those with a more profibrotic transcriptomic profile. SEMA3C expression is also elevated in different mouse models of liver fibrosis and in isolated HSCs on activation. In keeping with this, deletion of SEMA3C in activated HSCs reduces myofibroblast marker expression. Conversely, SEMA3C overexpression exacerbates TGF-ß-mediated myofibroblast activation, as shown by increased SMAD2 phosphorylation and target gene expression. Among SEMA3C receptors, only NRP2 expression is maintained on activation of isolated HSCs. Interestingly, lack of NRP2 in those cells reduces myofibroblast marker expression. Finally, deletion of either SEMA3C or NRP2, specifically in activated HSCs, reduces liver fibrosis in mice. CONCLUSION: SEMA3C is a novel marker for activated HSCs that plays a fundamental role in the acquisition of the myofibroblastic phenotype and liver fibrosis.

Development and Evaluation of MR-Based Radiogenomic Models to Differentiate Atypical Lipomatous Tumors from Lipomas.

BACKGROUND: The aim of this study was to develop and validate radiogenomic models to predict the MDM2 gene amplification status and differentiate between ALTs and lipomas on preoperative MR images. METHODS: MR images were obtained in 257 patients diagnosed with ALTs (n = 65) or lipomas (n = 192) using histology and the MDM2 gene analysis as a reference standard. The protocols included T2-, T1-, and fat-suppressed contrast-enhanced T1-weighted sequences. Additionally, 50 patients were obtained from a different hospital for external testing. Radiomic features were selected using mRMR. Using repeated nested cross-validation, the machine-learning models were trained on radiomic features and demographic information. For comparison, the external test set was evaluated by three radiology residents and one attending radiologist. RESULTS: A LASSO classifier trained on radiomic features from all sequences performed best, with an AUC of 0.88, 70% sensitivity, 81% specificity, and 76% accuracy. In comparison, the radiology residents achieved 60-70% accuracy, 55-80% sensitivity, and 63-77% specificity, while the attending radiologist achieved 90% accuracy, 96% sensitivity, and 87% specificity. CONCLUSION: A radiogenomic model combining features from multiple MR sequences showed the best performance in predicting the MDM2 gene amplification status. The model showed a higher accuracy compared to the radiology residents, though lower compared to the attending radiologist.

HAPLN1 potentiates peritoneal metastasis in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) frequently metastasizes into the peritoneum, which contributes to poor prognosis. Metastatic spreading is promoted by cancer cell plasticity, yet its regulation by the microenvironment is incompletely understood. Here, we show that the presence of hyaluronan and proteoglycan link protein-1 (HAPLN1) in the extracellular matrix enhances tumor cell plasticity and PDAC metastasis. Bioinformatic analysis showed that HAPLN1 expression is enriched in the basal PDAC subtype and associated with worse overall patient survival. In a mouse model for peritoneal carcinomatosis, HAPLN1-induced immunomodulation favors a more permissive microenvironment, which accelerates the peritoneal spread of tumor cells. Mechanistically, HAPLN1, via upregulation of tumor necrosis factor receptor 2 (TNFR2), promotes TNF-mediated upregulation of Hyaluronan (HA) production, facilitating EMT, stemness, invasion and immunomodulation. Extracellular HAPLN1 modifies cancer cells and fibroblasts, rendering them more immunomodulatory. As such, we identify HAPLN1 as a prognostic marker and as a driver for peritoneal metastasis in PDAC.

Outcome of patients with soft tissue sarcomas of the extremities and trunk treated by (neo)adjuvant intensity modulated radiation therapy with curative intent.

BACKGROUND: Soft tissue sarcomas (STS) are a relatively rare group of malignant tumors. Currently, there is very little published clinical data, especially in the context of curative multimodal therapy with image-guided, conformal, intensity-modulated radiotherapy. METHODS: Patients who received preoperative or postoperative intensity-modulated radiotherapy for STS of the extremities or trunk with curative intent were included in this single centre retrospective analysis. A Kaplan-Meier analysis was performed to evaluate survival endpoints. Multivariable proportional hazard models were used to investigate the association between survival endpoints and tumour-, patient-, and treatment-specific characteristics. RESULTS: 86 patients were included in the analysis. The most common histological subtypes were undifferentiated pleomorphic high-grade sarcoma (UPS) (27) and liposarcoma (22). More than two third of the patients received preoperative radiation therapy (72%). During the follow-up period, 39 patients (45%) suffered from some type of relapse, mainly remote (31%). The two-years overall survival rate was 88%. The median DFS was 48 months and the median DMFS was 51 months. Female gender (HR 0.460 (0.217; 0.973)) and histology of liposarcomas compared to UPS proved to be significantly more favorable in terms of DFS (HR 0.327 (0.126; 0.852)). CONCLUSION: Conformal, intensity-modulated radiotherapy is an effective treatment modality in the preoperative or postoperative management of STS. Especially for the prevention of distant metastases, the establishment of modern systemic therapies or multimodal therapy approaches is necessary.