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Introduction: The human leukocyte antigen complex (HLA) is essential for inducing specific immune responses to cancer by presenting tumor-associated peptides (TAP) to T cells. Overexpressed tumor associated antigens, mainly cancer-testis antigens (CTA), are outlined as essential targets for immunotherapy in oropharyngeal squamous cell carcinoma (OPSCC). This study assessed the degree to which presentation, gene expression, and antibody response (AR) of TAP, mainly CTA, are correlated in OPSCC patients to evaluate their potential as immunotherapy targets., Materials and Methods: Snap-frozen tumor (N Ligand/RNA =40), healthy mucosa (N RNA =6), and healthy tonsils (N Ligand =5) samples were obtained. RNA-Seq was performed using Illumina HiSeq 2500/NovaSeq 6000 and whole exome sequencing (WES) utilizing NextSeq500. HLA ligands were isolated from tumor tissue using immunoaffinity purification, UHPLC, and analyzed by tandem MS. Antibodies were measured in serum (N Ab =27) utilizing the KREX™ CT262 protein array. Data analysis focused on 312 proteins (KREX™ CT262 panel + overexpressed self-proteins)., Results: 183 and 94 of HLA class I and II TAP were identified by comparative profiling with healthy tonsils. Genes from 26 TAP were overexpressed in tumors compared to healthy mucosa (LFC>1; FDR<0.05). Low concordance (r=0.25; p<0.0001) was found between upregulated mRNA and class I TAP. The specific mode of correlation of TAP was found to be dependent on clinical parameters. A lack of correlation was observed both between mRNA and class II TAP, as well as between class II tumor-unique TAP (TAP-U) presentation and antibody response (AR) levels., Discussion: This study demonstrates that focusing exclusively on gene transcript levels fails to capture the full extent of TAP presentation in OPSCC. Furthermore, our findings reveal that although CTA are presented at relatively low levels, a few CTA TAP-U show potential as targets for immunotherapy., Competing Interests: SL: Advisory Boards: Merck Sharp & Dohme MSD, Bristol Myers Squibb BMS, Sanofi Genzyme, Astra Zeneca AZ. Honoraria: MSD, BMS. Travel reimbursement: Merck Serono, Astra Zeneca. The remaining 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. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Abou Kors, Meier, Mühlenbruch, Betzler, Oliveri, Bens, Thomas, Kraus, Doescher, von Witzleben, Hofmann, Ezic, Huber, Benckendorff, Barth, Greve, Schuler, Brunner, Blackburn, Hoffmann, Ottensmeier, Kestler, Rammensee, Walz and Laban.)

Background: Anti-vascular endothelial growth factor (VEGF) monoclonal antibodies (mAbs) are widely used for tumor treatment, including metastatic colorectal cancer (mCRC). So far, there are no biomarkers that reliably predict resistance to anti-VEGF mAbs like bevacizumab. A biomarker-guided strategy for early and accurate assessment of resistance could avoid the use of non-effective treatment and improve patient outcomes. We hypothesized that repeated analysis of multiple cytokines and angiogenic growth factors (CAFs) before and during treatment using machine learning could provide an accurate and earlier, i.e., 100 days before conventional radiologic staging, prediction of resistance to first-line mCRC treatment with FOLFOX plus bevacizumab., Patients and Methods: 15 German and Austrian centers prospectively recruited 50 mCRC patients receiving FOLFOX plus bevacizumab as first-line treatment. Plasma samples were collected every two weeks until radiologic progression (RECIST 1.1) as determined by CT scans performed every 2 months. 102 pre-selected CAFs were centrally analyzed using a cytokine multiplex assay (Luminex, Myriad RBM)., Results: Using random forests, we developed a predictive machine learning model that discriminated between the situations of "no progress within 100 days before radiological progress" and "progress within 100 days before radiological progress". We could further identify a combination of ten out of the 102 CAF markers, which fulfilled this task with 78.2% accuracy, 71.8% sensitivity, and 82.5% specificity., Conclusions: We identified a CAF marker combination that indicates treatment resistance to FOLFOX plus bevacizumab in patients with mCRC within 100 days prior to radiologic progress., Competing Interests: “•Thomas Seufferlein reports research funding to the institution from Sanofi during the conduct of the study; honoraria from Lilly, Pierre Fabre, BMS, MSD; advisory/consultancy roles with Amgen, Lilly, BMS, MSD, Pierre Fabre, Servier, Immodulon, Biontech, Mirati, Boehringer; Travel expenses from Takeda • Stefan Kasper discloses honoraria (self) from Merck, Amgen, Roche, Sanofi-Aventis, Servier, and Lilly; honoraria (institution) from Merck, Amgen, Roche, and Lilly; advisory/consultancy roles with Merck, BMS, Amgen, Roche, MSD, Sanofi-Aventis, Servier, and Lilly; research grants/funding (self) from Merck, Roche, BMS and Lilly; research grants/funding (institution) from Merck, Roche, BMS and Lilly; travel/accommodation expenses from Merck, Amgen, BMS, Roche, Sanofi, Aventis, Servier, and Lilly. • Thomas J. Ettrich reports honoraria from Roche, Sanofi; advisory/consultancy roles with Roche, Sanofi; research funding from Servier. • All other authors declare to have no conflicts of interest. This does not alter our adherence to PLOS ONE policies on sharing data and materials”., (Copyright: © 2024 Seufferlein et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

With the decline of bee populations worldwide, studies determining current wild bee distributions and diversity are increasingly important. Wild bee identification is often completed by experienced taxonomists or by genetic analysis. The current study was designed to compare two methods of identification including: (1) morphological identification by experienced taxonomists using images of field-collected wild bees and (2) genetic analysis of composite bee legs (multiple taxa) using metabarcoding. Bees were collected from conservation grasslands in eastern Iowa in summer 2019 and identified to the lowest taxonomic unit using both methods. Sanger sequencing of individual wild bee legs was used as a positive control for metabarcoding. Morphological identification of bees using images resulted in 36 unique taxa among 22 genera, and >80% of Bombus specimens were identified to species. Metabarcoding was limited to genus-level assignments among 18 genera but resolved some morphologically similar genera. Metabarcoding did not consistently detect all genera in the composite samples, including kleptoparasitic bees. Sanger sequencing showed similar presence or absence detection results as metabarcoding but provided species-level identifications for cryptic species (i.e., Lasioglossum). Genus-specific detections were more frequent with morphological identification than metabarcoding, but certain genera such as Ceratina and Halictus were identified equally well with metabarcoding and morphology. Genera with proportionately less tissue in a composite sample were less likely to be detected using metabarcoding. Image-based methods were limited by image quality and visible morphological features, while genetic methods were limited by databases, primers, and amplification at target loci. This study shows how an image-based identification method compares with genetic techniques, and how in combination, the methods provide valuable genus- and species-level information for wild bees while preserving tissue for other analyses. These methods could be improved and transferred to a field setting to advance our understanding of wild bee distributions and to expedite conservation research., Competing Interests: The authors have declared that no competing interests exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Patients with oropharyngeal squamous cell carcinoma (OPSCC) caused by human papilloma virus (HPV) exhibit a better prognosis than those with HPV-negative OPSCC. This study investigated the distinct molecular pathways that delineate HPV-negative from HPV-positive OPSCC to identify biologically relevant therapeutic targets. Bulk mRNA from 23 HPV-negative and 39 HPV-positive OPSCC tumors (n = 62) was sequenced to uncover the transcriptomic profiles. Differential expression followed by gene set enrichment analysis was performed to outline the top enriched biological process in the HPV-negative compared with HPV-positive entity. INHBA, the highest overexpressed gene in the HPV-negative tumor, was knocked down. Functional assays (migration, proliferation, cell death, stemness) were conducted to confirm the target's oncogenic role. Correlation analyses to reveal its impact on the tumor microenvironment were performed. We revealed that epithelial-to-mesenchymal transition (EMT) is the most enriched process in HPV-negative compared with HPV-positive OPSCC, with INHBA (inhibin beta A subunit) being the top upregulated gene. INHBA knockdown downregulated the expression of EMT transcription factors and attenuated migration, proliferation, stemness, and cell death resistance of OPSCC cells. We uncovered that INHBA associates with a pro-tumor microenvironment by negatively correlating with antitumor CD8+ T and B cells while positively correlating with pro-tumor M1 macrophages. We identified three miRNAs that are putatively involved in repressing INHBA expression. Our results indicate that the upregulation of INHBA is tumor-promoting. We propose INHBA as an attractive therapeutic target for the treatment of INHBA-enriched tumors in patients with HPV-negative OPSCC to ameliorate prognosis., Significance: Patients with HPV-negative OPSCC have a poorer prognosis due to distinct molecular pathways. This study reveals significant transcriptomic differences between HPV-negative and HPV-positive OPSCC, identifying INHBA as a key upregulated gene in HPV-negative OPSCC's oncogenic pathways. INHBA is crucial in promoting EMT, cell proliferation, and an immunosuppressive tumor environment, suggesting its potential as a therapeutic target for HPV-negative OPSCC., (© 2024 The Authors; Published by the American Association for Cancer Research.)

Pollinator diversity and abundance are declining globally. Cropland agriculture and the corresponding use of agricultural pesticides may contribute to these declines, while increased pollinator habitat (flowering plants) can help mitigate them. Here we tested whether the relative effect of wildflower plantings on pollinator diversity and counts were modified by proportion of nearby agricultural land cover and pesticide exposure in 24 conserved grasslands in Iowa, USA. Compared with general grassland conservation practices, wildflower plantings led to only a 5% increase in pollinator diversity and no change in counts regardless of the proportion of cropland agriculture within a 1 km radius. Pollinator diversity increased earlier in the growing season and with per cent flower cover. Unexpectedly, neither insecticide nor total pesticide concentrations on above-ground passive samplers were related to pollinator diversity. However, pollinator community composition was most strongly related to date of sampling, total pesticide concentration, and forb or flower cover. Our results indicate very little difference in pollinator diversity between grassland conservation practices with and without wildflower plantings. Given the relatively high economic costs of wildflower plantings, our research provides initial evidence that investment in general grassland conservation may efficiently conserve pollinator diversity in temperate regions of intensive cropland agriculture., Competing Interests: The work is all original research carried out by the authors. All authors agree with the contents of the manuscript and its submission to the journal. As explicitly described by the authors, the research featured in the manuscript relates to two other articles arising from the same larger study, one of which was published in Environmental Science and Technology and the other of which was recently submitted to PLoS ONE for review. All sources of funding are acknowledged above, and authors have no direct financial benefits that could result from publication to declare. All appropriate ethics and other approvals were obtained for the research., (© 2023 The Authors.)

Contamination of aquatic ecosystems can have cascading effects on terrestrial consumers by altering the availability and quality of aquatic insect prey. Comprehensive assessment of these indirect food-web effects of contaminants on natural resources and their associated services necessitates using both ecological and economic tools. In the present study we present an aquatic-terrestrial assessment tool (AT2), including ecological and economic decision trees, to aid practitioners and researchers in designing contaminant effect studies for linked aquatic-terrestrial insect-based food webs. The tool is tailored to address the development of legal claims by the US Department of the Interior's Natural Resource Damage Assessment and Restoration Program, which aims to restore natural resources injured by oil spills and hazardous substance releases into the environment. Such cases require establishing, through scientific inquiry, the existence of natural resource injury as well as the determination of the monetary or in-kind project-based damages required to restore this injury. However, this tool is also useful to researchers interested in questions involving the effects of contaminants on linked aquatic-terrestrial food webs. Stylized cases exemplify how application of AT2 can help practitioners and researchers design studies when the contaminants present at a site are likely to lead to injury of terrestrial aerial insectivores through loss of aquatic insect prey and/or dietary contaminant exposure. Designing such studies with ecological endpoints and economic modeling inputs in mind will increase the relevance and cost-effectiveness of studies, which can in turn improve the outcomes of cases and studies involving the ecological effects of contaminants on food webs. Environ Toxicol Chem 2023;42:2029-2039. Published 2023. This article is a U.S. Government work and is in the public domain in the USA. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC., (Published 2023. This article is a U.S. Government work and is in the public domain in the USA. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.)

Background: The immune peptidome of OPSCC has not previously been studied. Cancer-antigen specific vaccination may improve clinical outcome and efficacy of immune checkpoint inhibitors such as PD1/PD-L1 antibodies., Methods: Mapping of the OPSCC HLA ligandome was performed by mass spectrometry (MS) based analysis of naturally presented HLA ligands isolated from tumour tissue samples (n = 40) using immunoaffinity purification. The cohort included 22 HPV-positive (primarily HPV-16) and 18 HPV-negative samples. A benign reference dataset comprised of the HLA ligandomes of benign haematological and tissue datasets was used to identify tumour-associated antigens., Results: MS analysis led to the identification of naturally HLA-presented peptides in OPSCC tumour tissue. In total, 22,769 peptides from 9485 source proteins were detected on HLA class I. For HLA class II, 15,203 peptides from 4634 source proteins were discovered. By comparative profiling against the benign HLA ligandomic datasets, 29 OPSCC-associated HLA class I ligands covering 11 different HLA allotypes and nine HLA class II ligands were selected to create a peptide warehouse., Conclusion: Tumour-associated peptides are HLA-presented on the cell surfaces of OPSCCs. The established warehouse of OPSCC-associated peptides can be used for downstream immunogenicity testing and peptide-based immunotherapy in (semi)personalised strategies., (© 2023. The Author(s).)

Quantifying how contaminants change across life cycles of species that undergo metamorphosis is critical to assessing organismal risk, particularly for consumers. Pond-breeding amphibians can dominate aquatic animal biomass as larvae and are terrestrial prey as juveniles and adults. Thus, amphibians can be vectors of mercury exposure in both aquatic and terrestrial food webs. However, it is still unclear how mercury concentrations are affected by exogenous (e.g., habitat or diet) vs endogenous factors (e.g., catabolism during hibernation) as amphibians undergo large diet shifts and periods of fasting during ontogeny. We measured total mercury (THg), methylmercury (MeHg), and isotopic compositions (δ 13 C, δ 15 N) in boreal chorus frogs ( Pseudacris maculata ) across five life stages in two Colorado (USA) metapopulations. We found large differences in concentrations and percent MeHg (of THg) among life stages. Frog MeHg concentrations peaked during metamorphosis and hibernation coinciding with the most energetically demanding life cycle stages. Indeed, life history transitions involving periods of fasting coupled with high metabolic demands led to large increases in mercury concentrations. The endogenous processes of metamorphosis and hibernation resulted in MeHg bioamplification, thus decoupling it from the light isotopic proxies of diet and trophic position. These step changes are not often considered in conventional expectations of how MeHg concentrations within organisms are assessed.

Zebrafish (Danio rerio) are aquatic vertebrates with significant homology to their terrestrial counterparts. While zebrafish have a centuries-long track record in developmental and regenerative biology, their utility has grown exponentially with the onset of modern genetics. This is exemplified in studies focused on skeletal development and repair. Herein, the numerous contributions of zebrafish to our understanding of the basic science of cartilage, bone, tendon/ligament, and other skeletal tissues are described, with a particular focus on applications to development and regeneration. We summarize the genetic strengths that have made the zebrafish a powerful model to understand skeletal biology. We also highlight the large body of existing tools and techniques available to understand skeletal development and repair in the zebrafish and introduce emerging methods that will aid in novel discoveries in skeletal biology. Finally, we review the unique contributions of zebrafish to our understanding of regeneration and highlight diverse routes of repair in different contexts of injury. We conclude that zebrafish will continue to fill a niche of increasing breadth and depth in the study of basic cellular mechanisms of skeletal biology., (Copyright © 2022. Published by Elsevier Inc.)

Conservation efforts have been implemented in agroecosystems to enhance pollinator diversity by creating grassland habitat, but little is known about the exposure of bees to pesticides while foraging in these grassland fields. Pesticide exposure was assessed in 24 conservation grassland fields along an agricultural gradient at two time points (July and August) using silicone band passive samplers (nonlethal) and bee tissues (lethal). Overall, 46 pesticides were detected including 9 herbicides, 19 insecticides, 17 fungicides, and a plant growth regulator. For the bands, there were more frequent/higher concentrations of herbicides in July (maximum: 1600 ng/band in July; 570 ng/band in August), while insecticides and fungicides had more frequent/higher concentrations in August (maximum: 110 and 65 ng/band in July; 1500 and 1700 ng/band in August). Pesticide concentrations in bands increased 16% with every 10% increase in cultivated crops. The bee tissues showed no difference in detection frequency, and concentrations were similar among months; maximum concentrations of herbicides, insecticides, and fungicides in July and August were 17, 27, and 180 and 19, 120, and 170 ng/g, respectively. Pesticide residues in bands and bee tissues did not always show the same patterns; of the 20 compounds observed in both media, six (primarily fungicides) showed a detection-concentration relationship between the two media. Together, the band and bee residue data can provide a more complete understanding of pesticide exposure and accumulation in conserved grasslands.

Aim: Recently, a tumor cell-platelet interaction was identified in different tumor entities, resulting in a transfer of tumor-derived RNA into platelets, named further "tumor-educated platelets (TEP)". The present pilot study aims to investigate whether such a tumor-platelet transfer of RNA occurs also in patients suffering from head and neck squamous cell carcinoma (HNSCC)., Methods: Sequencing analysis of RNA derived from platelets of tumor patients (TPs) and healthy donors (HDs) were performed. Subsequently, quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used for verification of differentially expressed genes in platelets from TPs and HDs in a second cohort of patients and HDs. Data were analyzed by applying bioinformatic tools., Results: Sequencing of RNA derived from the tumor as well as from platelets of TPs and HDs revealed 426 significantly differentially existing RNA, at which 406 RNA were more and 20 RNA less abundant in platelets from TPs in comparison to that of HDs. In TPs' platelets, abundantly existing RNA coding for 49 genes were detected, characteristically expressed in epithelial cells and RNA, the products of which are involved in tumor progression. Applying bioinformatic tools and verification on a second TP/HD cohort, collagen type I alpha 1 chain (COL1A1) and zinc finger protein 750 (ZNF750) were identified as the strongest potentially platelet-RNA-sequencing (RNA-seq)-based biomarkers for HNSCC., Conclusions: These results indicate a transfer of tumor-derived messenger RNA (mRNA) into platelets of HNSCC patients. Therefore, analyses of a patient's platelet RNA could be an efficient option for liquid biopsy in order to diagnose HNSCC or to monitor tumorigenesis as well as therapeutic responses at any time and in real time., Competing Interests: The authors declare that they have no conflicts of interest., (© The Author(s) 2023.)

Historical mining left a legacy of abandoned mines and waste rock in remote headwaters of major river systems in the western United States. Understanding the influence of these legacy mines on culturally and ecologically important downstream ecosystems is not always straightforward because of elevated natural levels of mineralization in mining-impacted watersheds. To test the ecological effects of historic mining in the headwaters of the upper Salmon River watershed in Idaho (USA), we measured multiple community and chemical endpoints in downstream linked aquatic-terrestrial food webs. Mining inputs impacted downstream food webs through increased mercury accumulation and decreased insect biodiversity. Total mercury (THg) in seston, aquatic insect larvae, adult aquatic insects, riparian spiders, and fish at sites up to 7.6 km downstream of mining was found at much higher concentrations (1.3-11.3-fold) and was isotopically distinct compared with sites immediately upstream of mining inputs. Methylmercury concentrations in bull trout and riparian spiders were sufficiently high (732-918 and 347-1140 ng MeHg g -1  dry wt, respectively) to affect humans, birds, and piscivorous fish. Furthermore, the alpha-diversity of benthic insects was locally depressed by 12%-20% within 4.3-5.7 km downstream from the mine. However, because total insect biomass was not affected by mine inputs, the mass of mercury in benthic insects at a site (i.e., ng Hg m -2 ) was extremely elevated downstream (10-1778-fold) compared with directly upstream of mining inputs. Downstream adult aquatic insect-mediated fluxes of THg were also high (~16 ng THg m -2  day -1 ). Abandoned mines can have ecologically important effects on downstream communities, including reduced biodiversity and increased mercury flux to higher order consumers, including fish, birds, and humans. Environ Toxicol Chem 2022;41:1696-1710. Published 2022. This article is a U.S. Government work and is in the public domain in the USA., (Published 2022. This article is a U.S. Government work and is in the public domain in the USA.)

Controlling phenotypical landscapes is of vital interest to modern biology. This task becomes highly demanding because cellular decisions involve complex networks engaging in crosstalk interactions. Previous work on control theory indicates that small sets of compounds can control single phenotypes. However, a dynamic approach is missing to determine the drivers of the whole network dynamics. By analyzing 35 biologically motivated Boolean networks, we developed a method to identify small sets of compounds sufficient to decide on the entire phenotypical landscape. These compounds do not strictly prefer highly related compounds and show a smaller impact on the stability of the attractor landscape. The dynamic driver sets include many intervention targets and cellular reprogramming drivers in human networks. Finally, by using a new comprehensive model of colorectal cancer, we provide a complete workflow on how to implement our approach to shift from in silico to in vitro guided experiments., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Author(s).)

Metals and organic contaminants in aquatic systems affect the coupling of aquatic and terrestrial ecosystems through two pathways: contaminant-induced effects on insect emergence and emergence-induced contaminant transfer. Consequently, the impact of aquatic contaminants on terrestrial ecosystems can be driven by modifications in the quantity and quality of adult aquatic insects serving as prey or contaminants entering terrestrial food webs as part of the diet of terrestrial predators. Here, we provide an overview of recent advances in the field, separating metals from organic contaminants due to their differential propensity to bioaccumulate and thus their potential contribution to either of the two pathways. Finally, this review highlights the knowledge gap in the relative impact of these pathways on terrestrial insectivores., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Loss or damage to the mandible caused by trauma, treatment of oral malignancies, and other diseases is treated using bone-grafting techniques that suffer from numerous shortcomings and contraindications. Zebrafish naturally heal large injuries to mandibular bone, offering an opportunity to understand how to boost intrinsic healing potential. Using a novel her6:mCherry Notch reporter, we show that canonical Notch signaling is induced during the initial stages of cartilage callus formation in both mesenchymal cells and chondrocytes following surgical mandibulectomy. We also show that modulation of Notch signaling during the initial post-operative period results in lasting changes to regenerate bone quantity one month later. Pharmacological inhibition of Notch signaling reduces the size of the cartilage callus and delays its conversion into bone, resulting in non-union. Conversely, conditional transgenic activation of Notch signaling accelerates conversion of the cartilage callus into bone, improving bone healing. Given the conserved functions of this pathway in bone repair across vertebrates, we propose that targeted activation of Notch signaling during the early phases of bone healing in mammals may both augment the size of the initial callus and boost its ossification into reparative bone., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2022. Published by The Company of Biologists Ltd.)

The entorhinal cortex (EC) is the main interface between the sensory association areas of the neocortex and the hippocampus. It is crucial for the evaluation and processing of sensory data for long-term memory consolidation and shows damage in many brain diseases, for example, neurodegenerative diseases, such as Alzheimer's disease and developmental disorders. The pre-alpha layer of the EC in humans (layer II) displays a remarkable distribution of neurons in islands. These cellular islands give rise to a portion of the perforant path-the major reciprocal data stream for neocortical information into the hippocampal formation. However, the functional relevance of the morphological appearance of the pre-alpha layer in cellular islands and the precise timing of their initial appearance during primate evolution are largely unknown. Here, we conducted a comparative study of the EC from 38 nonhuman primates and Homo sapiens and found a strong relationship between gyrification index (GI) and the presence of the pre-alpha cellular islands. The formation of cellular islands also correlated with brain and body weight as well as neopallial volume. In the two human lissencephalic cases, the cellular islands in the pre-alpha layer were lacking. These findings emphasize the relationship between cortical folding and island formation in the EC from an evolutionary perspective and suggest a role in the pathomechanism of developmental brain disorders., (© 2021 The Authors. The Journal of Comparative Neurology published by Wiley Periodicals LLC.)

Pancreatic ductal adenocarcinomas (PDACs) are tumors with poor prognosis and limited treatment options. Personalized medicine aims at characterizing actionable DNA variants by next-generation sequencing, thereby improving treatment strategies and outcomes. Fine-needle tumor biopsies are currently the gold standard to acquire samples for DNA profiling. However, liquid biopsies have considerable advantages as they are minimally invasive and frequently obtainable and thus may help to monitor tumor evolution over time. However, which liquid analyte works best for this purpose is currently unclear. Our study aims to directly compare tumor-, circulating free (cf-) and extracellular vesicle-derived (ev)DNA by panel sequencing of matching patient material. We evaluated copy number variations (CNVs), single nucleotide variants (SNVs) and insertions and deletions (indels). Our data show that evDNA contains significantly larger DNA fragments up to 5.5 kb, in line with previous observations. Stringent bioinformatic processing revealed a significant advantage of evDNA with respect to cfDNA concerning detection performance for SNVs and a numerical increase for indels. A combination of ev- and cfDNA was clearly superior for SNV detection, as compared to either single analyte, thus potentially improving actionable variant prediction upon further optimization. Finally, calling of CNVs from liquid biopsies still remained challenging and uninformative.

Contaminants alter the quantity and quality of insect prey available to terrestrial insectivores. In agricultural regions, the quantity of aquatic insects emerging from freshwaters can be impacted by insecticides originating from surrounding croplands. We hypothesized that, in such regions, adult aquatic insects could also act as vectors of pesticide transfer to terrestrial food webs. To estimate insect-mediated pesticide flux from wetlands embedded in an important agricultural landscape, semipermanetly and temporarily ponded wetlands were surveyed in cropland and grassland landscapes across a natural salinity gradient in the Prairie Pothole Region of North Dakota (USA) during the bird breeding season in 2015 and 2016 (n = 14 and 15 wetlands, respectively). Current-use pesticides, including the herbicide atrazine and the insecticides bifenthrin and imidacloprid, were detected in newly emerged insects. Pesticide detections were similar in insects emerging from agricultural and grassland wetlands. Biomass of emerging aquatic insects decreased 43%, and insect-mediated pesticide flux increased 50% along the observed gradient in concentrations of insecticides in emerging aquatic insects (from 3 to 577 ng total insecticide g -1 insect). Overall, adult aquatic insects were estimated to transfer between 2 and 180 µg total pesticide wetland -1  d -1 to the terrestrial ecosystem. In one of the 2 study years, biomass of emerging adult aquatic insects was also 73% lower from agricultural than grassland wetlands and was dependent on salinity. Our results suggest that accumulated insecticides reduce the availability of adult aquatic insect prey for insectivores and potentially increase insectivore exposure to insect-borne pesticides. Adult aquatic insects retain pesticides across metamorphosis and may expose insectivores living near both agricultural and grassland wetlands to dietary sources of toxic chemicals. Environ Toxicol Chem 2021;40:2282-2296. Published 2021. This article is a U.S. Government work and is in the public domain in the USA., (Published 2021. This article is a U.S. Government work and is in the public domain in the USA.)

Background: Telomeres are protein-DNA complexes at the tips of linear chromosomes. They protect the DNA from end-to-end fusion and exonucleolytic degradation. Shortening of telomeric DNA during aging can generate dysfunctional telomeres, promoting tumorigenesis. More recent data indicate that both short and long telomeres of peripheral blood leukocyte (PBL) cells can serve as prognostic biomarkers for cancer risk and may be associated with survival of patients with solid cancers. Telomere length in PBL cells could also be a potential prognostic biomarker for survival in bladder cancer (BC) or renal cell carcinoma (RCC)., Methods: The relative telomere length (RTL) of PBL cells was assessed in patients with BC ( n = 144) and RCC ( n = 144) by using qPCR. A control population of patients without malignant disease (NC, n = 73) was included for comparison. The correlation and association of RTL with histopathological parameters and overall survival (OS) were evaluated., Results: Patients with BC and RCC had significantly shorter telomeres compared to patients without malignant disease. Within the cancer cohorts, multivariate analysis revealed that short RTL is an independent predictor of worse survival in BC ( p = 0.039) and RCC ( p = 0.041)., Conclusion: Patients with BC and RCC had significantly shorter telomeres compared to the normal population. Shorter RTL in BC and RCC was an independent predictor of reduced survival.

Background: Overcoming the COVID-19 crisis requires new ideas and strategies for online communication of personal medical information and patient empowerment. Rapid testing of a large number of subjects is essential for monitoring and delaying the spread of SARS-CoV-2 in order to mitigate the pandemic's consequences. People who do not know that they are infected may not stay in quarantine and, thus, risk infecting others. Unfortunately, the massive number of COVID-19 tests performed is challenging for both laboratories and the units that conduct throat swabs and communicate the results., Objective: The goal of this study was to reduce the communication burden for health care professionals. We developed a secure and easy-to-use tracking system to report COVID-19 test results online that is simple to understand for the tested subjects as soon as these results become available. Instead of personal calls, the system updates the status and the results of the tests automatically. This aims to reduce the delay when informing testees about their results and, consequently, to slow down the virus spread., Methods: The application in this study draws on an existing tracking tool. With this open-source and browser-based online tracking system, we aim to minimize the time required to inform the tested person and the testing units (eg, hospitals or the public health care system). The system can be integrated into the clinical workflow with very modest effort and avoids excessive load to telephone hotlines., Results: The test statuses and results are published on a secured webpage, enabling regular status checks by patients; status checks are performed without the use of smartphones, which has some importance, as smartphone usage diminishes with age. Stress tests and statistics show the performance of our software. CTest is currently running at two university hospitals in Germany-University Hospital Ulm and University Hospital Tübingen-with thousands of tests being performed each week. Results show a mean number of 10 (SD 2.8) views per testee., Conclusions: CTest runs independently of existing infrastructures, aims at straightforward integration, and aims for the safe transmission of information. The system is easy to use for testees. QR (Quick Response) code links allow for quick access to the test results. The mean number of views per entry indicates a reduced amount of time for both health care professionals and testees. The system is quite generic and can be extended and adapted to other communication tasks., (©Gunnar Völkel, Axel Fürstberger, Julian D Schwab, Silke D Werle, Nensi Ikonomi, Thomas Gscheidmeier, Johann M Kraus, Alexander Groß, Martin Holderried, Julien Balig, Franz Jobst, Peter Kuhn, Klaus A Kuhn, Oliver Kohlbacher, Udo X Kaisers, Thomas Seufferlein, Hans A Kestler. Originally published in the Journal of Medical Internet Research (https://www.jmir.org), 07.06.2021.)

The occurrence of adverse events frequently accompanies tumor treatments. Side effects should be detected and treated as soon as possible to maintain the best possible treatment outcome. Besides the standard reporting system Common Terminology Criteria for Adverse Events (CTCAE), physicians have recognized the potential of patient-reporting systems. These are based on a more subjective description of current patient reporting symptoms. Patient-reported symptoms are essential to define the impact of a given treatment on the quality of life and the patient's wellbeing. They also act against an underreporting of side effects which are paramount to define the actual value of a treatment for the individual patient. Here, we present a study protocol for a clinical trial that assesses the potential of a smartphone application for CTCAE conform symptom reporting and tracking that is adjusted to the standard clinical reporting system rather than symptom oriented descriptive trial tools. The presented study will be implemented in two parts, both lasting over six months. The first part will assess the feasibility of the application with 30 patients non-randomly divided into three equally-sized age groups (<55years, 55-75years, >75years). In the second part 36 other patients will be randomly assigned to two groups, one reporting using the smartphone and one not. This prospective second part will compare the impact of smartphone reported adverse events regarding applied therapy doses and quality of life to those of patients receiving standard care. We aim for early detection and treatment of adverse events in oncological treatment to improve patients' safety and outcomes. For this purpose, we will capture frequent adverse events of chemotherapies, immunotherapies, or other targeted therapies with our smartphone application. The presented trial is registered at the U.S. National Library of Medicine ClinicalTrials.gov (NCT04493450) on July 30, 2020., Competing Interests: The authors have declared that no competing interests exist.

Real-time isolation, propagation, and pharmacotyping of patient-derived pancreatic cancer organoids (PDOs) may enable treatment response prediction and personalization of pancreatic cancer (PC) therapy. In our methodology, PDOs are isolated from 54 patients with suspected or confirmed PC in the framework of a prospective feasibility trial. The drug response of single agents is determined by a viability assay. Areas under the curves (AUC) are clustered for each drug, and a prediction score is developed for combined regimens. Pharmacotyping profiles are obtained from 28 PDOs (efficacy 63.6%) after a median of 53 days (range 21-126 days). PDOs exhibit heterogeneous responses to the standard-of-care drugs, and are classified into high, intermediate, or low responder categories. Our developed prediction model allows a successful response prediction in treatment-naïve patients with an accuracy of 91.1% for first-line and 80.0% for second-line regimens, respectively. The power of prediction declines in pretreated patients (accuracy 40.0%), particularly with more than one prior line of chemotherapy. Progression-free survival (PFS) is significantly longer in previously treatment-naïve patients receiving a predicted tumor sensitive compared to a predicted tumor resistant regimen (mPFS 141 vs. 46 days; p = 0.0048). In conclusion, generation and pharmacotyping of PDOs is feasible in clinical routine and may provide substantial benefit.

Motivation: Cancer is a complex and heterogeneous disease involving multiple somatic mutations that accumulate during its progression. In the past years, the wide availability of genomic data from patients' samples opened new perspectives in the analysis of gene mutations and alterations. Hence, visualizing and further identifying genes mutated in massive sets of patients are nowadays a critical task that sheds light on more personalized intervention approaches., Results: Here, we extensively review existing tools for visualization and analysis of alteration data. We compare different approaches to study mutual exclusivity and sample coverage in large-scale omics data. We complement our review with the standalone software AVAtar ('analysis and visualization of alteration data') that integrates diverse aspects known from different tools into a comprehensive platform. AVAtar supplements customizable alteration plots by a multi-objective evolutionary algorithm for subset identification and provides an innovative and user-friendly interface for the evaluation of concurrent solutions. A use case from personalized medicine demonstrates its unique features showing an application on vaccination target selection., Availability: AVAtar is available at: https://github.com/sysbio-bioinf/avatar., Contact: hans.kestler@uni-ulm.de, phone: +49 (0) 731 500 24 500, fax: +49 (0) 731 500 24 502., (© The Author(s) 2020. Published by Oxford University Press.)

Aquatic insects link food web dynamics across freshwater-terrestrial boundaries and subsidize terrestrial consumer populations. Contaminants that accumulate in larval aquatic insects and are retained across metamorphosis can increase dietary exposure for riparian insectivores. To better understand potential exposure of terrestrial insectivores to aquatically-derived trace metals, metal concentrations in water and tissues were analyzed from different components of streams and riparian food webs across a large (2-3 orders of magnitude) metal gradient (e.g., Zn, Cu, Cd, Pb) in the Rocky Mountains (USA). Our research indicates that the trace metal concentration gradient present among streams was lost during metamorphosis of aquatic larval insects into terrestrially flying adults, decoupling terrestrial exposures from aquatic concentrations. This pattern was caused by declines in 1) among-stream variation in trace metal concentrations, 2) relationships between metal concentrations in paired water and food web components, and 3) mean metal concentrations within aquatic food webs and across the aquatic-terrestrial boundary. Specifically, among-stream variation in trace metal concentrations was highest for water and aquatic vegetation, intermediate for aquatic insect larvae (~30% lower than water) and lowest for adult aquatic insects and riparian spiders (~65% lower). Metal concentrations in paired water and food web components ranged from highly related across the stream-metal gradient (slopes ~1) for water and aquatic vegetation, to less related (slopes closer to 0) for aquatic vegetation and aquatic insect larvae, to unrelated (slopes ~0) for aquatic larval and adult insects. Finally, mean metal concentrations were highest in aquatic vegetation and lowest in adult aquatic insects emerging from streams (~50% lower than aquatic vegetation). Our results indicate less efficient trophic transfer and higher metamorphic loss of trace metals from high metal streams (i.e., exposure-dependent transfer). For many trace metals, aquatic-terrestrial dietary transfer is unlikely to be an important source of exposure for terrestrial insectivores of adult aquatic insects., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier B.V.)

Somatic cell reprogramming and tissue repair share relevant factors and molecular programs. Here, Dickkopf-3 (DKK3) is identified as novel factor for organ regeneration using combined transcription-factor-induced reprogramming and RNA-interference techniques. Loss of Dkk3 enhances the generation of induced pluripotent stem cells but does not affect de novo derivation of embryonic stem cells, three-germ-layer differentiation or colony formation capacity of liver and pancreatic organoids. However, DKK3 expression levels in wildtype animals and serum levels in human patients are elevated upon injury. Accordingly, Dkk3 -null mice display less liver damage upon acute and chronic failure mediated by increased proliferation in hepatocytes and LGR5 + liver progenitor cell population, respectively. Similarly, recovery from experimental pancreatitis is accelerated. Regeneration onset occurs in the acinar compartment accompanied by virtually abolished canonical-Wnt-signaling in Dkk3 -null animals. This results in reduced expression of the Hedgehog repressor Gli3 and increased Hedgehog-signaling activity upon Dkk3 loss. Collectively, these data reveal Dkk3 as a key regulator of organ regeneration via a direct, previously unacknowledged link between DKK3, canonical-Wnt-, and Hedgehog-signaling., Competing Interests: The authors declare no conflict of interest., (© 2021 The Authors. Advanced Science published by Wiley‐VCH GmbH.)

Programmed-death-1 (PD1) antibodies are approved for recurrent and metastatic head and neck squamous cell carcinoma. Multiple drugs targeting costimulatory and coinhibitory immune checkpoint molecules (ICM) have been discovered. However, it remains unknown how these ICM are affected by curative conventional therapy on different immune cell subsets during the course of treatment. In the prospective noninterventional clinical study titled "Immune Response Evaluation to Curative conventional Therapy" (NCT03053661), 22 patients were prospectively enrolled. Blood samples were drawn at defined time points throughout curative conventional treatment and follow-up. Immune cells (IC) from the different time points were assessed by multicolor flow cytometry. The following ICM were measured by flow cytometry: PD1, CTLA4, BTLA, CD137, CD27, GITR, OX40, LAG3 and TIM3. Dynamics of ICM expression were assessed using nonparametric paired samples tests. Significant changes were noted for PD1, BTLA and CD27 on multiple IC types during or after radiotherapy. Nonsignificant trends for increased expression of OX40 and GITR from baseline until the end of RT were observed on CD4 T cells and CD4 + CD39 + T cells. In patients with samples at recurrence of disease, a nonsignificant increase of TIM3 and LAG3 positive CD4 + CD39 + T cells was evident, accompanied by an increase of double positive cells for TIM3/LAG3. Potential future targets to be combined with RT in the conventional treatment and anti-PD1/PD-L could be BTLA agonists, or agonistic antibodies to costimulatory ICM like CD137, OX40 or GITR. The combination of cetuximab with CD27 agonistic antibodies enhancing ADCC or the targeting of TIM3/LAG3 may be another promising strategy., (© 2020 The Authors. International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.)

Objective: ATM serine/threonine kinase (ATM) is the most frequently mutated DNA damage response gene, involved in homologous recombination (HR), in pancreatic ductal adenocarcinoma (PDAC)., Design: Combinational synergy screening was performed to endeavour a genotype-tailored targeted therapy., Results: Synergy was found on inhibition of PARP, ATR and DNA-PKcs (PAD) leading to synthetic lethality in ATM-deficient murine and human PDAC. Mechanistically, PAD-induced PARP trapping, replication fork stalling and mitosis defects leading to P53-mediated apoptosis. Most importantly, chemical inhibition of ATM sensitises human PDAC cells toward PAD with long-term tumour control in vivo. Finally, we anticipated and elucidated PARP inhibitor resistance within the ATM-null background via whole exome sequencing. Arising cells were aneuploid, underwent epithelial-mesenchymal-transition and acquired multidrug resistance (MDR) due to upregulation of drug transporters and a bypass within the DNA repair machinery. These functional observations were mirrored in copy number variations affecting a region on chromosome 5 comprising several of the upregulated MDR genes. Using these findings, we ultimately propose alternative strategies to overcome the resistance., Conclusion: Analysis of the molecular susceptibilities triggered by ATM deficiency in PDAC allow elaboration of an efficient mutation-specific combinational therapeutic approach that can be also implemented in a genotype-independent manner by ATM inhibition., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

As service robots become increasingly autonomous and follow their own task-related goals, human-robot conflicts seem inevitable, especially in shared spaces. Goal conflicts can arise from simple trajectory planning to complex task prioritization. For successful human-robot goal-conflict resolution, humans and robots need to negotiate their goals and priorities. For this, the robot might be equipped with effective conflict resolution strategies to be assertive and effective but similarly accepted by the user. In this paper, conflict resolution strategies for service robots (public cleaning robot, home assistant robot) are developed by transferring psychological concepts (e.g., negotiation, cooperation) to HRI. Altogether, fifteen strategies were grouped by the expected affective outcome (positive, neutral, negative). In two online experiments, the acceptability of and compliance with these conflict resolution strategies were tested with humanoid and mechanic robots in two application contexts (public: n 1 = 61; private: n 2 = 93). To obtain a comparative value, the strategies were also applied by a human. As additional outcomes trust, fear, arousal, and valence, as well as perceived politeness of the agent were assessed. The positive/neutral strategies were found to be more acceptable and effective than negative strategies. Some negative strategies (i.e., threat, command) even led to reactance and fear. Some strategies were only positively evaluated and effective for certain agents (human or robot) or only acceptable in one of the two application contexts (i.e., approach, empathy). Influences on strategy acceptance and compliance in the public context could be found: acceptance was predicted by politeness and trust. Compliance was predicted by interpersonal power. Taken together, psychological conflict resolution strategies can be applied in HRI to enhance robot task effectiveness. If applied robot-specifically and context-sensitively they are accepted by the user. The contribution of this paper is twofold: conflict resolution strategies based on Human Factors and Social Psychology are introduced and empirically evaluated in two online studies for two application contexts. Influencing factors and requirements for the acceptance and effectiveness of robot assertiveness are discussed., 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 Babel, Kraus and Baumann.)

Background & Aims: Pancreatic tumor cells release small extracellular vesicles (sEVs, exosomes) that contain lipids and proteins, RNA, and DNA molecules that might promote formation of metastases. It is not clear what cargo these vesicles contain and how they are released. Protein kinase D1 (PRKD1) inhibits cell motility and is believed to be dysregulated in pancreatic ductal adenocarcinomas. We investigated whether it regulates production of sEVs in pancreatic cancer cells and their ability to form premetastatic niches for pancreatic cancer cells in mice., Methods: We analyzed data from UALCAN and human pancreatic tissue microarrays to compare levels of PRKD1 between tumor and nontumor tissues. We studied mice with pancreas-specific disruption of Prkd1 (PRKD1 KO mice), mice that express oncogenic KRAS (KC mice), and KC mice with disruption of Prkd1 (PRKD1 KO -KC mice). Subcutaneous xenograft tumors were grown in NSG mice from Panc1 cells; some mice were then given injections of sEVs. Pancreata and lung tissues from mice were analyzed by histology, immunohistochemistry, and/or quantitative polymerase chain reaction; we performed nanoparticle tracking analysis of plasma sEVs. The Prkd1 gene was disrupted in Panc1 cells using CRISPR-Cas9 or knocked down with small hairpin RNAs, or PRKD1 activity was inhibited with the selective inhibitor CRT0066101. Pancreatic cancer cell lines were analyzed by gene-expression microarray, quantitative polymerase chain reaction, immunoblot, and immunofluorescence analyses. sEVs secreted by Panc1 cell lines were analyzed by flow cytometry, transmission electron microscopy, and mass spectrometry., Results: Levels of PRKD1 were reduced in human pancreatic ductal adenocarcinoma tissues compared with nontumor tissues. PRKD1 KO -KC mice developed more pancreatic intraepithelial neoplasia, at a faster rate, than KC mice, and had more lung metastases and significantly shorter average survival time. Serum from PRKD1 KO -KC mice had increased levels of sEVs compared with KC mice. Pancreatic cancer cells with loss or inhibition of PRKD1 increased secretion of sEVs; loss of PRKD1 reduced phosphorylation of its substrate, cortactin, resulting in increased F-actin levels at the plasma membrane. sEVs from cells with loss or reduced expression of PRKD1 had altered content, and injection of these sEVs into mice increased metastasis of xenograft tumors to lung, compared with sEVs from pancreatic cells that expressed PRKD1. PRKD1-deficient pancreatic cancer cells showed increased loading of integrin α6β4 into sEVs-a process that required CD82., Conclusions: Human pancreatic ductal adenocarcinoma has reduced levels of PRKD1 compared with nontumor pancreatic tissues. Loss of PRKD1 results in reduced phosphorylation of cortactin in pancreatic cancer cell lines, resulting in increased in F-actin at the plasma membrane and increased release of sEVs, with altered content. These sEVs promote metastasis of xenograft and pancreatic tumors to lung in mice., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Second mitochondria-derived activator of caspase (SMAC) mimetics (SMs) targeting inhibitor of apoptosis proteins (IAPs) activate cell death pathways, and are currently being evaluated in clinical trials. Their successful therapeutic implementation requires upfront identification of patients who could benefit from a SM-based treatment but biomarkers for SM sensitivity have not yet been described. Here, we analyzed the intrinsic activity of two monovalent (AT406 and LCL161) and two bivalent (Birinapant and BV6) SMs on unselected patient-derived pediatric precursor B-cell acute lymphoblastic leukemia (BCP-ALL) identifying a subset of patient samples to be particularly sensitive to SM-induced cell death. This subset was defined by a characteristic gene expression signature with 127 differentially regulated genes, amongst them TNFRSF1A encoding TNFR1, and a critical role of TNFR1 in SM-induced cell death in sensitive BCP-ALL was confirmed on the functional level. Interestingly, samples with intermediate or low sensitivity to SMs were sensitized to SM-induced cell death by inhibition of caspases using zVAD.fmk or Emricasan, a pan-caspase inhibitor in clinical trials. When we compared our expression data to published data sets, we identified an overlap of four genes to be commonly differentially regulated in SM-sensitive BCP-ALL, that is, TSPAN7, DIPK1C, MTX2 and, again, TNFRSF1A. Functional testing revealed that this set of genes identified samples with high sensitivity to SM treatment. In summary, our data suggest using this gene signature as biomarker predicting response to SM treatment and point to the development of new combinatorial treatments consisting of SMs and pan-caspase inhibitors for a successful clinical implementation of SMs in treatment of BCP-ALL., (© 2019 The Authors. International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.)

Acute myeloid leukemia (AML) is the most common acute leukemia in adults and is propagated by leukemic stem cells (LSCs), often characterized by deregulated Wnt signaling. We previously showed that the central transcriptional mediator of Wnt signaling LEF1 is able to cause AML in mice and acts as an independent prognostic factor in normal karyotype AML. Here, we show that treatment naïve normal karyotype AML as well as samples AML LSCs predominantly express the long β-catenin-binding isoform of LEF1 in sharp contrast to normal human hematopoietic stem cells, which lack expression of the long isoform, but express the short N-terminally truncated isoform with loss of the β-catenin-binding site. Gene expression and ChiP-Seq analyses in mice linked the long isoform to Wnt-β-catenin signaling and oncogenic pathways, the N-terminally truncated isoform to stemness associated genes. Approaches impairing binding of LEF1 to β-catenin significantly impaired AML growth, but spared normal hematopoietic stem cells. This report now demonstrates a striking difference of LEF1 isoform expression between normal and AML cells, contributing to higher vulnerability of leukemic cells to approaches targeting β-catenin/LEF1 interaction.

Accumulation of DNA damage and myeloid-skewed differentiation characterize aging of the hematopoietic system, yet underlying mechanisms remain incompletely understood. Here, we show that aging hematopoietic progenitor cells particularly of the myeloid branch exhibit enhanced resistance to bulky DNA lesions-a relevant type of DNA damage induced by toxins such as cancer drugs or endogenous aldehydes. We identified aging-associated activation of the Hedgehog (Hh) pathway to be connected to this phenotype. Inhibition of Hh signaling reverts DNA damage tolerance and DNA damage-resistant proliferation in aged hematopoietic progenitors. Vice versa, elevating Hh activity in young hematopoietic progenitors is sufficient to impair DNA damage responses. Altogether, these findings provide experimental evidence for aging-associated increases in Hh activity driving DNA damage tolerance in myeloid progenitors and myeloid-skewed differentiation. Modulation of Hh activity could thus be explored as a therapeutic strategy to prevent DNA damage tolerance, myeloid skewing, and disease development in the aging hematopoietic system.

There have been hints that nonviral cancer antigens are differentially expressed in human papillomavirus (HPV)-positive and HPV-negative head and neck squamous cell carcinoma (HNSCC). Antibody responses (AR) to cancer antigens may be used to indirectly determine cancer antigen expression in the tumor using a noninvasive and tissue-saving liquid biopsy. Here, we set out to characterize AR to a panel of nonviral cancer antigens in HPV-positive and HPV-negative HNSCC patients. A fluorescent microbead multiplex serology to 29 cancer antigens (16 cancer-testis antigens, 5 cancer-retina antigens and 8 oncogenes) and 29 HPV-antigens was performed in 382 HNSCC patients from five independent cohorts (153 HPV-positive and 209 HPV-negative). AR to any of the cancer antigens were found in 272/382 patients (72%). The ten most frequent AR were CT47, cTAGE5a, c-myc, LAGE-1, MAGE-A1, -A3, -A4, NY-ESO-1, SpanX-a1 and p53. AR to MAGE-A3, MAGE-A9 and p53 were found at significantly different prevalences by HPV status. An analysis of AR mean fluorescent intensity values uncovered remarkably different AR clusters by HPV status. To identify optimal antigen selections covering a maximum of patients with ≤10 AR, multiobjective optimization revealed distinct antigen selections by HPV status. We identified that AR to nonviral antigens differ by HPV status indicating differential antigen expression. Multiplex serology may be used to characterize antigen expression using serum or plasma as a tissue-sparing liquid biopsy. Cancer antigen panels should address the distinct antigen repertoire of HPV-positive and HPV-negative HNSCC., (© 2019 The Authors. International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.)

Purpose: The identification of high-risk patients within human papillomavirus (HPV)-positive and -negative head and neck squamous cell carcinoma (HNSCC) is needed for improved treatment and surveillance strategies. In this study, we set out to discover antibody responses (AR) with prognostic impact in HNSCC stratified by HPV status., Experimental Design: A fluorescent bead-based multiplex serology assay on 29 cancer antigens (16 cancer-testis antigens, 5 cancer-retina antigens, and 8 oncogenes) and 29 HPV antigens was performed in samples of 362 patients with HNSCC from five independent cohorts (153 HPV positive, 209 HPV negative). A multivariable Cox proportional hazard model with bootstrapping (M = 1000) was used for validation of prognostic antibody responses., Results: Antibody response to any of the cancer antigens was found in 257 of 362 patients (71%). In HPV-negative patients, antibody responses to c-myc, MAGE-A1, -A4, and Rhodopsin E2 (combined as AR high risk ) were significantly associated with shorter overall survival. In HPV-positive patients, antibody responses to IMP-1 were discovered as a negative prognostic factor. AR high risk (HR = 1.76) and antibody responses to IMP-1 (HR = 3.28) were confirmed as independent markers for a poor prognosis in a multivariable Cox proportional hazard model with bootstrapping (M = 1000)., Conclusions: We identified antibody responses to cancer antigens that associate with a dismal prognosis in patients with HNSCC beyond HPV-positive status. AR high risk may be used to detect HPV-negative patients with an extraordinarily bad prognosis. Most importantly, antibody response to IMP-1 may serve as a marker for a subgroup of HPV-positive patients who present with a poor prognosis similar to that in HPV-negative patients., (©2019 American Association for Cancer Research.)

Deregulated cell death pathways contribute to leukemogenesis and treatment failure in B-cell precursor acute lymphoblastic leukemia (BCP-ALL). Intrinsic apoptosis signaling is regulated by different proapoptotic and antiapoptotic molecules: proapoptotic BCL-2 homology domain 3 (BH3) proteins activate prodeath molecules leading to cellular death, while antiapoptotic molecules including B-cell lymphoma 2 (BCL-2) prevent activation of prodeath proteins and counter-regulate apoptosis induction. Inhibition of these antiapoptotic regulators has become a promising strategy for anticancer treatment, but variable anticancer activities in different malignancies indicate the need for upfront identification of responsive patients. Here, we investigated the activity of the BCL-2 inhibitor venetoclax (VEN, ABT-199) in B-cell precursor acute lymphoblastic leukemia and found heterogeneous sensitivities in BCP-ALL cell lines and in a series of patient-derived primografts. To identify parameters of sensitivity and resistance, we evaluated genetic aberrations, gene-expression profiles, expression levels of apoptosis regulators, and functional apoptosis parameters analyzed by mitochondrial profiling using recombinant BH3-like peptides. Importantly, ex vivo VEN sensitivity was most accurately associated with functional BCL-2 dependence detected by BH3 profiling. Modeling clinical application of VEN in a preclinical trial in a set of individual ALL primografts, we identified that leukemia-free survival of VEN treated mice was precisely determined by functional BCL-2 dependence. Moreover, the predictive value of ex vivo measured functional BCL-2 dependence for preclinical in vivo VEN response was confirmed in an independent set of primograft ALL including T- and high risk-ALL. Thus, integrative analysis of the apoptosis signaling indicating mitochondrial addiction to BCL-2 accurately predicts antileukemia activity of VEN, robustly identifies VEN-responsive patients, and provides information for stratification and clinical guidance in future clinical applications of VEN in patients with ALL.

Dynamic models analyzing gene regulation and metabolism face challenges when adapted to modeling signal transduction networks. During signal transduction, molecular reactions and mechanisms occur in different spatial and temporal frames and involve feedbacks. This impedes the straight-forward use of methods based on Boolean networks, Bayesian approaches, and differential equations. We propose a new approach, ProbRules, that combines probabilities and logical rules to represent the dynamics of a system across multiple scales. We demonstrate that ProbRules models can represent various network motifs of biological systems. As an example of a comprehensive model of signal transduction, we provide a Wnt network that shows remarkable robustness under a range of phenotypical and pathological conditions. Its simulation allows the clarification of controversially discussed molecular mechanisms of Wnt signaling by predicting wet-lab measurements. ProbRules provides an avenue in current computational modeling by enabling systems biologists to integrate vast amounts of available data on different scales., Competing Interests: The authors declare no competing interests.

Organism aging is characterized by increased inflammation and decreased stem cell function, yet the relationship between these factors remains incompletely understood. This study shows that aged hematopoietic stem and progenitor cells (HSPCs) exhibit increased ground-stage NF-κB activity, which enhances their responsiveness to undergo differentiation and loss of self-renewal in response to inflammation. The study identifies Rad21 /cohesin as a critical mediator of NF-κB signaling, which increases chromatin accessibility in the vicinity of NF-κB target genes in response to inflammation. Rad21 is required for normal differentiation, but limits self-renewal of hematopoietic stem cells (HSCs) during aging and inflammation in an NF-κB-dependent manner. HSCs from aged mice fail to down-regulate Rad21 /cohesin and inflammation/differentiation signals in the resolution phase of inflammation. Inhibition of cohesin/NF-κB reverts hypersensitivity of aged HSPCs to inflammation-induced differentiation and myeloid-biased HSCs with disrupted/reduced expression of Rad21 /cohesin are increasingly selected during aging. Together, Rad21 /cohesin-mediated NF-κB signaling limits HSPC function during aging and selects for cohesin-deficient HSCs with myeloid-skewed differentiation., (© 2018 Chen et al.)

Liver cholestasis is a chronic liver disease and a major health problem worldwide. Cholestasis is characterised by a decrease in bile flow due to impaired secretion by hepatocytes or by obstruction of bile flow through intra- or extrahepatic bile ducts. Thereby cholestasis can induce ductal proliferation, hepatocyte injury and liver fibrosis. Notch signalling promotes the formation and maturation of bile duct structures. Here we investigated the liver regeneration process in the context of cholestasis induced by disruption of the Notch signalling pathway. Liver-specific deletion of recombination signal binding protein for immunoglobulin kappa j region ( Rbpj ), which represents a key regulator of Notch signalling, induces severe cholestasis through impaired intra-hepatic bile duct (IHBD) maturation, severe necrosis and increased lethality. Deregulation of the biliary compartment and cholestasis are associated with the change of several signalling pathways including a Kyoto Encyclopedia of Genes and Genomes (KEGG) gene set representing the Hippo pathway, further yes-associated protein (YAP) activation and upregulation of SRY (sex determining region Y)-box 9 (SOX9), which is associated with transdifferentiation of hepatocytes. SOX9 upregulation in cholestatic liver injury in vitro is independent of Notch signalling. We could comprehensively address that in vivo Rbpj depletion is followed by YAP activation, which influences the transdifferentiation of hepatocytes and thereby contributing to liver regeneration.

Dysregulated intestinal epithelial apoptosis initiates gut injury, alters the intestinal barrier, and can facilitate bacterial translocation leading to a systemic inflammatory response syndrome (SIRS) and/or multi-organ dysfunction syndrome (MODS). A variety of gastrointestinal disorders, including inflammatory bowel disease, have been linked to intestinal apoptosis. Similarly, intestinal hyperpermeability and gut failure occur in critically ill patients, putting the gut at the center of SIRS pathology. Regulation of apoptosis and immune-modulatory functions have been ascribed to Thirty-eight-negative kinase 1 (TNK1), whose activity is regulated merely by expression. We investigated the effect of TNK1 on intestinal integrity and its role in MODS. TNK1 expression induced crypt-specific apoptosis, leading to bacterial translocation, subsequent septic shock, and early death. Mechanistically, TNK1 expression in vivo resulted in STAT3 phosphorylation, nuclear translocation of p65, and release of IL-6 and TNF-α. A TNF-α neutralizing antibody partially blocked development of intestinal damage. Conversely, gut-specific deletion of TNK1 protected the intestinal mucosa from experimental colitis and prevented cytokine release in the gut. Finally, TNK1 was found to be deregulated in the gut in murine and porcine trauma models and human inflammatory bowel disease. Thus, TNK1 might be a target during MODS to prevent damage in several organs, notably the gut.

We investigated polychlorinated biphenyl (PCB) contamination at the Ashtabula River (northeast OH, USA) area of concern following remedial dredging using araneid and tetragnathid spiders. The PCB concentrations remain elevated in the area of concern compared with reference conditions. Patterns of contamination were strikingly similar between taxa, but were higher in tetragnathids at the most contaminated sites. Spider PCB homolog distributions identified 2 PCB sources to the area of concern. Based on these findings, we recommend situations where these taxa can be used alone, in concert, or combined into a composite "spider" sample to assess environmental contamination. Environ Toxicol Chem 2018;37:2467-2474. Published 2018 Wiley Periodicals Inc. on behalf of SETAC. This article is a US government work and, as such, is in the public domain in the United States of America., (Published 2018 Wiley Periodicals Inc. on behalf of SETAC. This article is a US government work and, as such, is in the public domain in the United States of America.)

Diversity and biomass of aquatic insects decline in metal-rich aquatic environments, but the mechanisms by which insects from such environments cope with potentially toxic metal concentrations to survive through adulthood are less well understood. In this study, we measured Zn concentrations and isotopes in laboratory-reared diatoms and mayflies (Neocloeon triangulifer) from larval through adult stages. The larvae were fed Zn-enriched diatoms, and bio-concentrated Zn by a factor of 2.5-5 relative to the diatoms but maintained the same Zn-isotopic ratio. These results reflect the importance of dietary uptake and the greater rate of uptake relative to excretion or growth. Upon metamorphosis to subimago, Zn concentrations declined by >70%, but isotopically heavy Zn remained in the subimago bodies. We surmised that the loss of isotopically light Zn during metamorphosis was due to the loss of detoxified Zn and retention of metabolically useful Zn. Through the transition from subimago to imago, Zn concentrations and isotope ratios were virtually unchanged. Because the decrease in Zn body concentration and increase in heavier Zn are seen in the subimagos relative to the larvae, the compartmentalization of Zn must be occurring within the larvae. A biodynamic model was constructed, allowing for isotopic fractionation and partitioning of Zn between metabolically essential and detoxified Zn reservoirs within larvae. The model provides a consistent set of rate and fractionation constants that successfully describe the experimental observations. Specifically, metabolically essential Zn is isotopically heavier and is tightly held once assimilated, and excess, isotopically light Zn is sequestered, detoxified, and ultimately lost during the metamorphosis of larvae to subimagos. To our knowledge, this is the first documentation of in vivo isotopic fractionation in insects, offering an improved understanding of the mechanisms and rates by which the N. triangulifer larvae regulate excess Zn in their bodies., (Published by Elsevier B.V.)