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Immune activation status determines non-small cell lung cancer (NSCLC) prognosis, with reported positive/negative associations for T helper type 2 (TH2) responses, including allergen-specific IgE and eosinophils. Our study seeks to explore the potential impact of these comorbid immune responses on the survival rates of patients with NSCLC. Our retrospective study used data from the Data Warehouse of the German Center for Lung Research (DZL) and Lung Biobank at Thoraxklinik Heidelberg. We estimated the association of blood eosinophilia and neutrophilia on survival rates in an inflammatory cohort of 3143 patients with NSCLC. We also tested sensitization to food and inhalants and high-sensitivity C-reactive protein (hs-CRP) in a comorbidity cohort of 212 patients with NSCLC. Finally, we estimated the infiltration of immune-relevant cells including eosinophils, T-cells, and mast cells in a tissue inflammatory sub-cohort of 60 patients with NSCLC. Sensitization to at least one food or inhalant (sIgE) was higher in patients with adenocarcinoma (adeno-LC) than the non-adenocarcinoma (non-adeno-LC). Furthermore, hs-CRP was higher in non-adeno-LC compared with adeno-LC. Peripheral inflammation, particularly eosinophilia and neutrophilia, was associated with poor survival outcomes in NSCLC with a clear difference between histological subgroups. Finally, blood eosinophilia was paralleled by significant eosinophil infiltration into the peritumoral tissue in the lung. This study provides novel perspectives on the crucial role of peripheral inflammation, featuring eosinophilia and neutrophilia, with overall survival, underscoring distinctions between NSCLC subgroups (adeno-LC vs. non-adeno-LC). Peripheral eosinophilia enhances eosinophil infiltration into tumors. This sheds light on the complex interplay between inflammation, eosinophil infiltration, and NSCLC prognosis among various histological subtypes. Further studies are required to underscore the role of eosinophils in NSCLC among different histological subgroups and their role in shaping the tumor microenvironment.

Neuroendocrine tumors of the small intestine (SI-NETs) often develop lymph node metastasis (LNM)-induced mesenteric fibrosis (MF). MF can cause intestinal obstruction as well as ischemia and render surgical resection technically challenging. The underlying pathomechanisms of MF are still not well understood. We examined mesenteric LNM and the surrounding stroma compartment from 24 SI-NET patients, including 11 with in situ presentation of strong MF (MF+) and 13 without MF (MF-). Differential gene expression was assessed with the HTG EdgeSeq Oncology Biomarker Panel comparing MF+ with MF- within LNM and paired stromal samples, respectively. Most interesting differentially expressed genes were validated by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) in combination with validation of associated protein levels utilizing immunohistochemistry (IHC) staining of MF+ and MF- formalin-fixed, paraffin-embedded (FFPE) patient samples. Overall, 14 genes measured with a 2549-gene expression panel were differentially expressed in MF+ patients compared to MF-. Of those, nine were differentially expressed genes in LNM and five genes in the stromal tissue (>2-fold change, p < .05). The top hits included increased COMP and COL11A1 expression in the stroma of MF+ patients compared to MF-, as well as decreased HMGA2, COL6A6, and SLC22A3 expression in LNM of MF+ patients compared to LNM of MF- patients. RT-qPCR confirmed high levels of COMP and COL11A1 in stroma samples of MF+ compared to MF- patients. IHC staining confirmed the enrichment of α-smooth muscle actin-positive fibrosis in MF+ compared to MF- patients with corresponding increase of COMP-expressing stromal cells in MF+. Since COMP is associated with the known driver for fibrosis development transforming growth factor beta and with a cancer-associated fibroblasts enriched environment, it seems to be a promising new target for MF research., (© 2024 The Authors. Journal of Neuroendocrinology published by John Wiley & Sons Ltd on behalf of British Society for Neuroendocrinology.)

A crucial requirement for metastasis formation in ovarian high-grade serous carcinoma (HGSC) is the disruption of the protective peritoneal mesothelium. Using co-culture systems of primary human cells, we discovered that tumor-associated NK cells induce TRAIL-dependent apoptosis in mesothelial cells via death receptors DR4 and DR5 upon encounter with activated T cells. Upregulation of TRAIL expression in NK cells concomitant with enhanced cytotoxicity toward mesothelial cells was driven predominantly by T-cell-derived TNFα, as shown by affinity proteomics-based analysis of the T cell secretome in conjunction with functional studies. Consistent with these findings, we detected apoptotic mesothelial cells in the peritoneal fluid of HGSC patients. In contrast to mesothelial cells, HGSC cells express negligible levels of both DR4 and DR5 and are TRAIL resistant, indicating cell-type-selective killing by NK cells. Our data point to a cooperative action of T and NK in breaching the mesothelial barrier in HGSC patients., Competing Interests: The authors declare no competing interests., (© 2023 The Author(s).)

Intratumoral cytotoxic CD8+ T cells (CTL) enter a dysfunctional state characterized by expression of coinhibitory receptors, loss of effector function, and changes in the transcriptional landscape. Even though several regulators of T-cell exhaustion have been identified, the molecular mechanisms inducing T-cell exhaustion remain unclear. Here, we show that IL18 receptor (IL18R) signaling induces CD8+ T-cell exhaustion in a murine pancreatic cancer model. Adoptive transfer of Il18r-/- OT-1 CD8+ CTLs resulted in enhanced rejection of subcutaneous tumors expressing ovalbumin (OVA) as a model antigen (PancOVA), compared with wild-type OT-1 CTLs. Transferred intratumoral IL18R-deficient CTLs expressed higher levels of effector cytokines TNF and IFNγ and had reduced expression of coinhibitory receptors (PD-1, TIM-3, 2B4, LAG-3) and the transcription factors Eomes and TOX. Lower expression of coinhibitory receptors and TOX on IL18R-deficient versus IL18R-sufficient CD8+ T cells were confirmed in an orthotopic KPC model. IL18R-induced T-cell exhaustion was regulated by IL2/STAT5 and AKT/mTOR pathways, as demonstrated in an in vitro exhaustion assay. Concordantly, mice deficient in NLRP3, the molecular complex activating IL18, had decreased expression of coinhibitory receptors on intratumoral T cells and similar changes in signaling pathways at the transcriptome level. Thus, molecular pathways promoting T-cell exhaustion indicate an involvement of an NLRP3-expressing tumor microenvironment, which mediates IL18 release. The Cancer Genome Atlas analysis of patients with pancreatic carcinoma showed an association between NLRP3-mediated IL18 signaling and shorter survival. These findings indicate NLRP3-mediated IL18R signaling as a regulator of intratumoral T-cell exhaustion and a possible target for immunotherapy. See related Spotlight by Stromnes, p. 400., (©2023 American Association for Cancer Research.)

Background: Toll-like receptors (TLRs) are gaining attention for their potential to influence tumor biology both on the level of the tumor cells as well as on the level of the surrounding inflammatory stroma. Previous studies resulted in partly conflicting data on the expression of TLR7 in healthy and neoplastic pancreatic tissues as well as its role in pancreatic tumor biology., Methods: We used qRT-PCR and immunohistochemistry to asses TLR7 expression in primary patient material and cell lines. Cell viability was analyzed by MTT assay upon incubation with TLR7 agonist/antagonist. Mouse models were used to investigate the role of TLR7 in vivo., Results: TLR7 is overexpressed in more than 50% of primary human pancreatic ductal adenocarcinoma (PDAC). High TLR7 expression was associated with shorter patient survival, and TLR7 inhibition in cell lines reduced viability in a dose-dependent manner. In contrast, global TLR7 deficiency did not alter survival or overall histopathological tumor features in genetic mouse models of PDAC., Conclusions: TLR7 may have opposing functions in tumor versus stroma cells. Further work is required to more precisely dissect the roles of TLR7 and its ligands in different populations of epithelial and stromal cells and to understand their relative contributions to tumor progression., (© 2023 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

The abnormal tumor microenvironment (TME) often dictates the therapeutic response of cancer to chemo- and immuno-therapy. Aberrant expression of pericentromeric satellite repeats has been reported for epithelial cancers, including lung cancer. However, the transcription of tandemly repetitive elements in stromal cells of the TME has been unappreciated, limiting the optimal use of satellite transcripts as biomarkers or anti-cancer targets. We found that transcription of pericentromeric satellite DNA (satDNA) in mouse and human lung adenocarcinoma was observed in cancer-associated fibroblasts (CAFs). In vivo, lung fibroblasts expressed pericentromeric satellite repeats HS2/HS3 specifically in tumors. In vitro, transcription of satDNA was induced in lung fibroblasts in response to TGFβ, IL1α, matrix stiffness, direct contact with tumor cells and treatment with chemotherapeutic drugs. Single-cell transcriptome analysis of human lung adenocarcinoma confirmed that CAFs were the cell type with the highest number of satellite transcripts. Human HS2/HS3 pericentromeric transcripts were detected in the nucleus, cytoplasm, extracellularly and co-localized with extracellular vesicles in situ in human biopsies and activated fibroblasts in vitro. The transcripts were transmitted into recipient cells and entered their nuclei. Knock-down of satellite transcripts in human lung fibroblasts attenuated cellular senescence and blocked the formation of an inflammatory CAFs phenotype which resulted in the inhibition of their pro-tumorigenic functions. In sum, our data suggest that satellite long non-coding (lnc) RNAs are induced in CAFs, regulate expression of inflammatory genes and can be secreted from the cells, which potentially might present a new element of cell-cell communication in the TME., (© 2023. The Author(s).)

Immune checkpoint blockade therapy is a treatment option of various metastatic cancer diseases including renal cell carcinoma (RCC). Approved antibody drugs target the co-inhibitory signaling of Programmed Cell Death Ligand-1 (PD-L1) and its receptor Programmed Cell Death-1 (PD-1). The combined evaluation of PD-L1 and PD-1 at the mRNA and protein levels in tumor tissue with differentiation of tumor and immune cells as well as of soluble forms (sPD-L1) and (sPD-1) in blood is of basic interest in assessing biomarker surrogates. Here, we demonstrate that PD-L1 determined as fraction of stained tumor cells (TPS-score) correlates with PD-L1-mRNA in tumor tissue, reflecting the predominant expression of PD-L1 in tumor cells. Conversely, PD-1 in immune cells of tumor tissue (IC-score) correlated with PD-1-mRNA tissue levels reflecting the typical PD-1 expression in immune cells. Of note, sPD-L1 in blood did not correlate with either the TPS-score of PD-L1 or with PD-L1-mRNA in tumor tissue. sPD-L1 released into the supernatant of cultured RCC cells closely followed the cellular PD-L1 expression as tested by interferon γ (IFNG) induction and siRNA knockdown of PD-L1. Further analysis in patients revealed that sPD-L1 significantly increased in blood following renal tumor resection. In addition, sPD-L1 correlated significantly with inflammation marker C-reactive protein (CRP) and with PD-L1 mRNA level in whole blood. These results indicate that the major source of sPD-L1 in blood may be peripheral blood cells and not primarily tumor tissue PD-L1., (© 2022. The Author(s).)

Background: The intestinal microbiota fundamentally guides the development of a normal intestinal physiology, the education, and functioning of the mucosal immune system. The Citrobacter rodentium-carrier model in germ-free (GF) mice is suitable to study the influence of selected microbes on an otherwise blunted immune response in the absence of intestinal commensals., Results: Here, we describe that colonization of adult carrier mice with 14 selected commensal microbes (OMM 12 + MC 2 ) was sufficient to reestablish the host immune response to enteric pathogens; this conversion was facilitated by maturation and activation of the intestinal blood vessel system and the step- and timewise stimulation of innate and adaptive immunity. While the immature colon of C. rodentium-infected GF mice did not allow sufficient extravasation of neutrophils into the gut lumen, colonization with OMM 12 + MC 2 commensals initiated the expansion and activation of the visceral vascular system enabling granulocyte transmigration into the gut lumen for effective pathogen elimination., Conclusions: Consortium modeling revealed that the addition of two facultative anaerobes to the OMM 12 community was essential to further progress the intestinal development. Moreover, this study demonstrates the therapeutic value of a defined consortium to promote intestinal maturation and immunity even in adult organisms. Video Abstract., (© 2022. The Author(s).)

Chloromas, also referred to as myeloid sarcomas, describe rare extramedullary tumor aggregates of malignant myeloid progenitor cells. The aim of this study was investigate the diagnostic features and characteristics of chloromas using contrast-enhanced ultrasound (CEUS). Between July 2007 and April 2021, 15 patients with 20 myeloid neoplasms and suspected chloroma manifestations were examined using B-mode US (B-US) and CEUS. Clinical data and B-US (echogenicity, border, size) and CEUS (hyper-, iso-, hypo- or complex enhancement) characteristics were retrospectively analyzed. Absolute and relative frequencies were determined. In B-US, the chloromas were most frequently hypo-echoic (n = 15, 75%). In addition, a hyperechoic (n = 2, 10%) or echocomplex (n = 3, 15%) presentation was observed. On CEUS, 7 chloromas (35%) had an arterial hyperenhancement, 8 (40%) an iso-enhancement and 3 (15%) a complex enhancement. Two chloromas (10%) did not exhibit any enhancement. We describe for the first time CEUS and B-US patterns of chloromas. They are typically hypo-echoic on B-US and have a strong iso- or hyperenhancement on CEUS, which may help in the differential diagnosis of some unclear masses (e.g., hematoma, abscess) in patients with myeloid neoplasias. Nevertheless, histology is necessary for a reliable diagnosis., (Copyright © 2022 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.)

Background: Impaired p53 function is one of the central molecular features of a tumor cell and even a partial reduction in p53 activity can increase the cancer risk in mice and men. From a therapeutic perspective it is noteworthy that tumor cells often become addicted to the absence of p53 providing a rationale for developing p53 reactivating compounds to treat cancer patients. Unfortunately, many of the compounds that are currently undergoing preclinical and clinical testing fail to fully reactivate mutant p53 proteins, raising the crucial question: how much p53 activity is needed to elicit a therapeutic effect?, Methods: We have genetically modelled partial p53 reactivation using knock-in mice with inducible expression of the p53 variant E177R. This variant has a reduced ability to bind and transactivate target genes and consequently causes moderate cancer susceptibility. We have generated different syngeneically transplanted and autochthonous mouse models of p53-deficient acute myeloid leukemia and B or T cell lymphoma. After cancer manifestation we have activated E177R expression and analyzed the in vivo therapy response by bioluminescence or magnetic resonance imaging. The molecular response was further characterized in vitro by assays for gene expression, proliferation, senescence, differentiation, apoptosis and clonogenic growth., Results: We report the conceptually intriguing observation that the p53 variant E177R, which promotes de novo leukemia and lymphoma formation, inhibits proliferation and viability, induces immune cell infiltration and triggers cancer regression in vivo when introduced into p53-deficient leukemia and lymphomas. p53-deficient cancer cells proved to be so addicted to the absence of p53 that even the low-level activity of E177R is detrimental to cancer growth., Conclusions: The observation that a partial loss-of-function p53 variant promotes tumorigenesis in one setting and induces regression in another, underlines the highly context-specific effects of individual p53 mutants. It further highlights the exquisite sensitivity of cancer cells to even small changes in p53 activity and reveals that changes in activity level are more important than the absolute level. As such, the study encourages ongoing research efforts into mutant p53 reactivating drugs by providing genetic proof-of-principle evidence that incomplete p53 reactivation may suffice to elicit a therapeutic response., (© 2022. The Author(s).)

Purpose: To describe the perfusion patterns of peripheral pulmonary granulomatous lesions (PPGLs) by contrast-enhanced ultrasound (CEUS) and their correlation with vascularization patterns (VPs) represented by immunohistochemical (CD34) endothelial staining., Patients and Methods: From January 2007 until September 2020, 10 consecutive patients with histologically confirmed PPGLs were investigated by CEUS. The time to enhancement, classified as early pulmonary-arterial (PA) pattern of enhancement versus delayed bronchial-arterial (BA) pattern of enhancement, the extent of enhancement, classified as marked or reduced, the homogeneity of enhancement, classified as homogeneous or inhomogeneous, and the decrease of enhancement, classified as rapid washout (<120 seconds) or a late washout (≥120 seconds), were analyzed retrospectively. Furthermore, the tissue samples from the study patients and as a control group, 10 samples of normal lung tissue obtained by autopsy, and 10 samples of lung tissue with acute pneumonia obtained by autopsy were immunohistochemically stained with CD34 antibody. The presence of avascular areas (AAs) and the VPs were evaluated in all tissue samples., Results: On CEUS, all PPGLs showed a reduced inhomogeneous BA pattern of enhancement and a rapid washout (<120 seconds). On CD34 staining, all PPGLs showed central AAs in granulomas and a chaotic VP similar to angiogenesis in lung tumors. The lung tissue in control groups revealed on CD34 staining a regular alveolar VP., Conclusion: The PPGLs on CEUS show an identical perfusion pattern similar to those of malignant lesions. Furthermore, for the first time, neoangiogenesis was demonstrated as a histopathological correlate to BA pattern of enhancement on CEUS., (© 2021 The Authors. Journal of Ultrasound in Medicine published by Wiley Periodicals LLC on behalf of American Institute of Ultrasound in Medicine.)

Purpose: To describe perfusion patterns of peripheral pulmonary lesions (PPLs) in COVID-19 patients using contrast-enhanced ultrasound (CEUS)., Patients and Methods: From April 2020 until July 2020, 11 consecutive patients with RT-PCR-confirmed COVID-19 and PPLs sized over 5 mm were investigated by B-mode ultrasound (B-US) and CEUS. The homogeneity of enhancement (homogeneous and inhomogeneous) was examined retrospectively using CEUS. An inhomogeneous enhancement was defined as a perfused lesion with coexisting non-perfused areas (NPA)., Results: On B-US, all 11 patients showed an interstitial syndrome (B-lines) with PPLs between 0.5 and 6 cm. On CEUS, all cases showed peripheral NPA during the complete CEUS examination. One patient underwent a partial lung resection with subsequent histopathological examination. The histological examination showed vasculitis, microthrombus in the alveolar capillary, and small obliterated vessels., Conclusion: In our case series, PPLs in patients with RT-PCR-confirmed COVID-19 infection presented a CEUS pattern with NPA during the complete CEUS examination. Our findings suggest a peripheral pulmonary perfusion disturbance in patients with COVID-19 infection. In 1 case, the histopathological correlation with the perfusion disturbance in the PPL was proven., (© 2021 The Authors. Journal of Ultrasound in Medicine published by Wiley Periodicals LLC on behalf of American Institute of Ultrasound in Medicine.)

Purpose: To describe the perfusion patterns of peripheral organizing pneumonia (POP) by contrast-enhanced ultrasound (CEUS) and their correlation with vascularization patterns (VPs) represented by immunohistochemical CD34 endothelial staining. Methods: From October 2006 until December 2020, 38 consecutive patients with histologically confirmed POPs were standardized-examined by CEUS. The time to enhancement (TE; classified as an early pulmonary-arterial [PA] pattern of enhancement vs. delayed bronchial-arterial [BA] pattern of enhancement), the extent of enhancement (EE; classified as marked or reduced), the homogeneity of enhancement (HE; classified as homogeneous or inhomogeneous), and the decrease of enhancement (DE; classified as rapid washout [<120s] or late washout [≥120s]) were evaluated retrospectively. Furthermore, tissue samples from the study patients were immunohistochemically stained with CD34 antibody. The presence of avascular areas (AAs) and the VPs were evaluated in all tissue samples. Results: The majority of POPs showed a BA pattern of enhancement (71.1%), an isoechoic marked enhancement (76.3%), and an inhomogeneous enhancement (81.6%). A rapid DE was observed in 50.0% of cases. On CD34 staining, all POPs had a chaotic VP, indicating BA neoangiogenesis. AAs (abscess, necrosis, hemorrhage) were identified in (41.9%) cases with an inhomogeneous enhancement on CEUS. Conclusion: On CEUS, POPs predominantly revealed a marked inhomogeneous BA pattern of enhancement with a rapid washout in 50% of cases. Furthermore, we demonstrated that the presence of a PA pattern of enhancement, found in 28.9% of POPs, did not exclude a BA neoangiogenesis as an important feature of chronic inflammatory and malignant processes.

The p14 ARF protein is a well-known regulator of p53-dependent and p53-independent tumor-suppressive activities. In unstressed cells, p14 ARF is predominantly sequestered in the nucleoli, bound to its nucleolar interaction partner NPM. Upon genotoxic stress, p14 ARF undergoes an immediate redistribution to the nucleo- and cytoplasm, where it promotes activation of cell cycle arrest and apoptosis. Here, we identify p14 ARF as a novel interaction partner and substrate of PRMT1 (protein arginine methyltransferase 1). PRMT1 methylates several arginine residues in the C-terminal nuclear/nucleolar localization sequence (NLS/NoLS) of p14 ARF . In the absence of cellular stress, these arginines are crucial for nucleolar localization of p14 ARF . Genotoxic stress causes augmented interaction between PRMT1 and p14 ARF , accompanied by arginine methylation of p14 ARF . PRMT1-dependent NLS/NoLS methylation promotes the release of p14 ARF from NPM and nucleolar sequestration, subsequently leading to p53-independent apoptosis. This PRMT1-p14 ARF cooperation is cancer-relevant and indicative for PDAC (pancreatic ductal adenocarcinoma) prognosis and chemotherapy response of pancreatic tumor cells. Our data reveal that PRMT1-mediated arginine methylation is an important trigger for p14 ARF 's stress-induced tumor-suppressive function., (© 2021 The Authors. Published under the terms of the CC BY NC ND 4.0 license.)

Postulated by Strachan more than 30 years ago, the Hygiene Hypothesis has undergone many revisions and adaptations. This review journeys back to the beginnings of the Hygiene Hypothesis and describes the most important landmarks in its development considering the many aspects that have refined and generalized the Hygiene Hypothesis over time. From an epidemiological perspective, the Hygiene Hypothesis advanced to a comprehensive concept expanding beyond the initial focus on allergies. The Hygiene Hypothesis comprise immunological, microbiological and evolutionary aspects. Thus, the original postulate developed into a holistic model that explains the impact of post-modern life-style on humans, who initially evolved in close proximity to a more natural environment. Focusing on diet and the microbiome as the most prominent exogenous influences we describe these discrepancies and the resulting health outcomes and point to potential solutions to reestablish the immunological homeostasis that frequently have been lost in people living in developed societies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Pfefferle, Keber, Cohen and Garn.)