Analyse und Funktionalität von αβ-T-Zellen im klinischen Kontext einer Patienten-individuellen anti-Tumor-Vakzinierung und der Graft-versus-Host-Disease

Hämatologische Erkrankungen wie Leukämien sowie solide Tumore mit Metastasenbildung haben häufig eine schlechte Prognose. Standardisierte Therapien, wie Chemo- und Strahlentherapie, können initial Remissionen erzeugen sind aber häufig mit einem Rezidiv verbunden und in Folgetherapien weniger wirksam, da die Tumorzellen einen erworbenen Selektionsvorteil gegen diese Therapie entwickelt haben. Die Immuntherapie wird immer wichtiger in der Tumortherapie. Basierend auf der Physiologie der Immunzellen kann spezifisch mit dem eigenen Immunsystem oder nach hämatopoetischer Stammzellentransplantation (HSCT) mit allogenen Immunzellen der Tumor/ die Leukämie bekämpft werden. Eine Form der Therapie ist die anti-Tumor-Vakzinierung. Aus Tumorgewebe werden die Mutationen mit Next-Generation Sequencing (NGS) identifiziert, entsprechende Impfpeptide designt und diese, analog zu den klassischen Impfungen, dem Patienten in Kombination mit einem Adjuvans appliziert und so die T-Zellen des Patienten gegen den eigenen Tumor gerichtet. Eine weitere Form der Immuntherapie, besonders bei Leukämiepatienten angewandt, ist die HSCT. In Vorbereitung auf die Transplantation werden die Patienten aufgrund der maximalen Chemotherapie immundefizient, da die hoch konzentrierte Chemotherapeutika alle hämatopoetischen Zellen zerstört. Nach anschließender allogener HSCT bilden transplantierte Stammzellen, nach dem Anwachsen, der Differenzierung und Proliferation in dem Patienten, ein gesundes Immunsystem. Dieses reagiert im optimalen Fall gegen die Leukämiezellen. Beide Therapien, die HSCT sowie die Patienten-individuelle anti-Tumor-Vakzinierung nutzen das Potenzial aktivierter T Zellen und können dem Patienten mit Krebs einen Überlebensvorteil ermöglichen. Im Gegensatz zu den erwünschten T-Zellantworten sind aktivierte T-Zellen, die nach einer HSCT, das Gewebe und die Organe des Empfängers angreifen und eine Autoimmunerkrankung oder eine graft-versus-host disease (GvHD) induzieren unerwünscht. Therapien mit Kortison und Immunsuppressiva führen häufig zu lebensbedrohlichen Infektionen und reduzieren den Behandlungserfolg. Neue Therapiestrategien für Abstoßungsreaktionen fehlen und Forschungen bezüglich neuer Medikamente sind aufgrund fehlender Tiermodelle mit humaner chronischer GvHD schwierig. Für den ersten Teil der Arbeit, die Analyse immunreaktiver T-Zellen nach individueller anti-Tumor-Vakzinierung untersuchten wir, bei einem, an Pankreaskarzinom erkrankten 62-jährigen Patienten, nach Resektion und Detektion zweier Missense -Mutationen in einer Lebermetastase, über Exomsequenzierung, ob die darauf abgestimmten vier synthetisierten Impfpeptide mit Tumor-spezifischer Mutation, unter Berücksichtigung der MHC-Restriktion des Patienten, eine anti-tumorale T-Zellantwort induzieren konnten. Die Impfung erfolgte monatlich in einem Zeitraum von 5 Jahren. Die durch Impfpeptide reaktivierten T-Zellen, isolierten und untersuchten wir an 3 verschiedenen Zeitpunkten, nach 12, 17 und 34 Impfungen und konnten eine erfolgreich etablierte anti-tumorale Impfpeptid- und T-Zell-spezifische Immunantwort nachweisen. Basierend auf den molekular identifizierten CDR3-Regionen dieser T Zellen konnten wir sowohl Persistenz und Breite der induzierten Immunantwort zeigen sowie durch konvergente CDR3-Regionen der TCRs Rückschlüsse auf die Immunogenität der Impfpetide schließen. Der Patient profitierte von der Vakzinierung, er zeigte keine Nebenwirkungen und lebte 6 Jahre ohne Rezidiv. Im zweiten Teil der Arbeit, untersuchten wir 15 CD34+ selektionierte / CD3+ depletierte humane Stammzelltransplantate in NSG Mäusen auf ihr Potenzial eine Abstoßung zu induzieren. Wir konnten zeigen, dass transplantierte CD34+ humane Stammzellen in den Mäusen anwachsen und in 2 der 15 Transplantaten ab der 24. Woche sich eine systemische, autoimmune cGvHD entwickelte, die in Lunge, Leber, Haut, Gingiva und Darm auftrat. Die beiden Transplantate unterschieden sich von den Transplantaten der anderen 13 Spendern in den HLA-Merkmalen, das heißt beide GvHD induzierende Transplantate zeigten einen Haplotyp, der mit einem erhöhten Risiko für die Entwicklung einer GvHD oder Autoimmunerkrankung assoziiert ist. Beide Haplotypen werden definiert als HASEH engl.: human autoimmunity susceptibility extended HLA haplotypes. Die reaktiven T-Zellen, CD4+ TH2-Zellen, mit autoreaktiven CDR3-Regionen, exprimieren in den betroffenen Geweben der erkrankten Mäuse, verstärkt die profibrotischen Zytokine IL-13 und IL-4 und verursachen sclerodermatöse Haut, eine fibrotische Lunge, verschiedene Hepatitis und massive Zahndeformationen bis hin zum Zahnverlust. Die Besonderheit dieses CD34+ NSG-Modells, weder Thymus oder Thymus-abgeleitete regulatorische T-Zellen (nTreg) noch B-Zellen konnten nachgewiesen werden, was die Rolle des genetischen Polymorphismus und der Zytokine bezüglich der Pathogenese der cGvHD verdeutlicht. Die Etablierung eines solchen Modells, das erstmals die chronische, autoimmune Variante der Abstoßungsreaktion in seiner multiorganen Pathologie induziert, eignet sich für Untersuchungen, die zur Aufklärung der Pathophysiologie humaner Autoimmunität und cGvHD beitragen und bietet die Möglichkeit, neue Behandlungsstrategien zu entwickeln und zu untersuchen

Decreased feeding rates of the copepod Acartia tonsa when exposed to playback harbor traffic noise

Copepods present the largest and most diverse group of zooplankton and their feeding behavior can affect top-down and bottom-up processes. Thus, how efficient feeding is executed determines the abundance of copepods’ prey and their predators and, with that, carbon transfer and storage in ecosystems. The rise of anthropogenic underwater noise from shipping, oil exploration and exploitation, wind farm construction and operation, and more, is increasingly changing the marine acoustic environment. This acoustic pollution can have detrimental effects on biological life. Studies on this topic increasingly indicate that anthropogenic underwater noise adversely affects primary producers, marine mammals, fish, and invertebrates. However, little data exist on the effects of anthropogenic underwater noise on the feeding behavior of zooplankton.We measured significantly decreased ingestion rates and clearance rates of A. tonsa when exposed to harbor noise compared to ambient conditions. The negative impact of noise on the ingestion rates was found at all given phytoplankton cell densities between 1k to 10k cells ml−1. Clearance rates were fitted to the Rogers random predator equation which revealed significantly decreased capture rates on phytoplankton under the exposure of harbor noise while handling times remained the same in both sound treatments.Our results call for follow-up studies to focus on noise driven community-effects in field experiments to confirm laboratory results and to predict the outcome of a changing world with multiple stressors. Further, the underlying mechanism on how noise affects the feeding behavior of copepods is still unknown. Noise may distract copepods or mask hydromechanical cues of the prey. Noise may also adversely affect copepod physiology or morphology that would lead to changes in the feeding behavior. All potential mechanisms need to be investigated rigorously in future experiments

Religiöser Wandel in muslimischen Gesellschaften

Processes of individualization in Egypt, Iran, Lebanon, Senegal and Tunisia are central to current social dynamics and form the key theme of the present volume. The central question is how social transformations inform the field of the religious and how processes of religious change likewise have an impact on social change. The contributions in this volume focus in particular on the realities of life for women from urban middle-class milieus. They show how women’s efforts to shape one's individual life may run counter to established social and religious norms. While analyzing milieu-specific dynamics, particular attention is devoted to the question how processes of social and religious change become manifest in both gender and generational relations. Western ideals of modernity, globalized discourses and respective leisure/fashion-practices as well as expressions of individualized lifestyles have an impact on dynamics of social and religious change and are discussed in this volume in specific religious as well as non-religious social contexts

Pseudomonas aeruginosa lectin LecB inhibits tissue repair processes by triggering β-catenin degradation

AbstractPseudomonas aeruginosa is an opportunistic pathogen that induces severe lung infections such as ventilator-associated pneumonia and acute lung injury. Under these conditions, the bacterium diminishes epithelial integrity and inhibits tissue repair mechanisms, leading to persistent infections. Understanding the involved bacterial virulence factors and their mode of action is essential for the development of new therapeutic approaches.In our study we discovered a so far unknown effect of the P. aeruginosa lectin LecB on host cell physiology. LecB alone was sufficient to attenuate migration and proliferation of human lung epithelial cells and to induce transcriptional activity of NF-κB. These effects are characteristic of impaired tissue repair. Moreover, we found a strong degradation of β-catenin, which was partially recovered by the proteasome inhibitor lactacystin. In addition, LecB induced loss of cell–cell contacts and reduced expression of the β-catenin targets c-myc and cyclin D1. Blocking of LecB binding to host cell plasma membrane receptors by soluble l-fucose prevented these changes in host cell behavior and signaling, and thereby provides a powerful strategy to suppress LecB function.Our findings suggest that P. aeruginosa employs LecB as a virulence factor to induce β-catenin degradation, which then represses processes that are directly linked to tissue recovery

Religiöser Wandel in muslimischen Gesellschaften

Processes of individualization in Egypt, Iran, Lebanon, Senegal and Tunisia are central to current social dynamics and form the key theme of the present volume. The central question is how social transformations inform the field of the religious and how processes of religious change likewise have an impact on social change. The contributions in this volume focus in particular on the realities of life for women from urban middle-class milieus. They show how women’s efforts to shape one's individual life may run counter to established social and religious norms. While analyzing milieu-specific dynamics, particular attention is devoted to the question how processes of social and religious change become manifest in both gender and generational relations. Western ideals of modernity, globalized discourses and respective leisure/fashion-practices as well as expressions of individualized lifestyles have an impact on dynamics of social and religious change and are discussed in this volume in specific religious as well as non-religious social contexts

Comparing ion conductance recordings of synthetic lipid bilayers with cell membranes containing TRP channels

In this article we compare electrical conductance events from single channel recordings of three TRP channel proteins (TRPA1, TRPM2 and TRPM8) expressed in human embryonic kidney cells with channel events recorded on synthetic lipid membranes close to melting transitions. Ion channels from the TRP family are involved in a variety of sensory processes including thermo- and mechano-reception. Synthetic lipid membranes close to phase transitions display channel-like events that respond to stimuli related to changes in intensive thermodynamic variables such as pressure and temperature. TRP channel activity is characterized by typical patterns of current events dependent on the type of protein expressed. Synthetic lipid bilayers show a wide spectrum of electrical phenomena that are considered typical for the activity of protein ion channels. We find unitary currents, burst behavior, flickering, multistep-conductances, and spikes behavior in both preparations. Moreover, we report conductances and lifetimes for lipid channels as described for protein channels. Non-linear and asymmetric current-voltage relationships are seen in both systems. Without further knowledge of the recording conditions, no easy decision can be made whether short current traces originate from a channel protein or from a pure lipid membraneComment: 13 pages, 9 Figure

EIF2AK2 Missense Variants Associated with Early Onset Generalized Dystonia

Objective: The study was undertaken to identify a monogenic cause of early onset, generalized dystonia. Methods: Methods consisted of genome-wide linkage analysis, exome and Sanger sequencing, clinical neurological examination, brain magnetic resonance imaging, and protein expression studies in skin fibroblasts from patients. Results: We identified a heterozygous variant, c.388G&gt;A, p.Gly130Arg, in the eukaryotic translation initiation factor 2 alpha kinase 2 (EIF2AK2) gene, segregating with early onset isolated generalized dystonia in 5 patients of a Taiwanese family. EIF2AK2 sequencing in 191 unrelated patients with unexplained dystonia yielded 2 unrelated Caucasian patients with an identical heterozygous c.388G&gt;A, p.Gly130Arg variant, occurring de novo in one case, another patient carrying a different heterozygous variant, c.413G&gt;C, p.Gly138Ala, and one last patient, born from consanguineous parents, carrying a third, homozygous variant c.95A&gt;C, p.Asn32Thr. These 3 missense variants are absent from gnomAD, and are located in functional domains of the encoded protein. In 3 patients, additional neurological manifestations were present, including intellectual disability and spasticity. EIF2AK2 encodes a kinase (protein kinase R [PKR]) that phosphorylates eukaryotic translation initiation factor 2 alpha (eIF2α), which orchestrates the cellular stress response. Our expression studies showed abnormally enhanced activation of the cellular stress response, monitored by PKR-mediated phosphorylation of eIF2α, in fibroblasts from patients with EIF2AK2 variants. Intriguingly, PKR can also be regulated by PRKRA (protein interferon-inducible double-stranded RNA-dependent protein kinase activator A), the product of another gene causing monogenic dystonia. Interpretation: We identified EIF2AK2 variants implicated in early onset generalized dystonia, which can be dominantly or recessively inherited, or occur de novo. Our findings provide direct evidence for a key role of a dysfunctional eIF2α pathway in the pathogenesis of dystonia. ANN NEUROL 2021;89:485–497.</p

LIPAD (LRRK2/Luebeck International Parkinson's Disease) Study Protocol:Deep Phenotyping of an International Genetic Cohort

Background: Pathogenic variants in the Leucine-rich repeat kinase 2 (LRRK2) gene are the most common known monogenic cause of Parkinson's disease (PD). LRRK2-linked PD is clinically indistinguishable from idiopathic PD and inherited in an autosomal dominant fashion with reduced penetrance and variable expressivity that differ across ethnicities and geographic regions.Objective: To systematically assess clinical signs and symptoms including non-motor features, comorbidities, medication and environmental factors in PD patients, unaffected LRRK2 pathogenic variant carriers, and controls. A further focus is to enable the investigation of modifiers of penetrance and expressivity of LRRK2 pathogenic variants using genetic and environmental data.Methods: Eligible participants are invited for a personal or online examination which comprises completion of a detailed eCRF and collection of blood samples (to obtain DNA, RNA, serum/plasma, immune cells), urine as well as household dust. We plan to enroll 1,000 participants internationally: 300 with LRRK2-linked PD, 200 with LRRK2 pathogenic variants but without PD, 100 PD patients with pathogenic variants in the GBA or PRKN genes, 200 patients with idiopathic PD, and 200 healthy persons without pathogenic variants.Results: The eCRF consists of an investigator-rated (1 h) and a self-rated (1.5 h) part. The first part includes the Movement Disorder Society Unified Parkinson's Disease Rating, Hoehn &amp;Yahr, and Schwab &amp; England Scales, the Brief Smell Identification Test, and Montreal Cognitive Assessment. The self-rating part consists of a PD risk factor, food frequency, autonomic dysfunction, and quality of life questionnaires, the Pittsburgh Sleep Quality Inventory, and the Epworth Sleepiness as well as the Hospital Anxiety and Depression Scales. The first 15 centers have been initiated and the first 150 participants enrolled (as of March 25th, 2021).Conclusions: LIPAD is a large-scale international scientific effort focusing on deep phenotyping of LRRK2-linked PD and healthy pathogenic variant carriers, including the comparison with additional relatively frequent genetic forms of PD, with a future perspective to identify genetic and environmental modifiers of penetrance and expressivityClinical Trial Registration:ClinicalTrials.gov, NCT04214509

Large-Scale Screening: Phenotypic and Mutational Spectrum in Isolated and Combined Dystonia Genes

© 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.[Background] Pathogenic variants in several genes have been linked to genetic forms of isolated or combined dystonia. The phenotypic and genetic spectrum and the frequency of pathogenic variants in these genes have not yet been fully elucidated, neither in patients with dystonia nor with other, sometimes co-occurring movement disorders such as Parkinson's disease (PD).[Objectives] To screen >2000 patients with dystonia or PD for rare variants in known dystonia-causing genes.[Methods] We screened 1207 dystonia patients from Germany (DysTract consortium), Spain, and South Korea, and 1036 PD patients from Germany for pathogenic variants using a next-generation sequencing gene panel. The impact on DNA methylation of KMT2B variants was evaluated by analyzing the gene's characteristic episignature.[Results] We identified 171 carriers (109 with dystonia [9.0%]; 62 with PD [6.0%]) of 131 rare variants (minor allele frequency <0.005). A total of 52 patients (48 dystonia [4.0%]; four PD [0.4%, all with GCH1 variants]) carried 33 different (likely) pathogenic variants, of which 17 were not previously reported. Pathogenic biallelic variants in PRKRA were not found. Episignature analysis of 48 KMT2B variants revealed that only two of these should be considered (likely) pathogenic.[Conclusion] This study confirms pathogenic variants in GCH1, GNAL, KMT2B, SGCE, THAP1, and TOR1A as relevant causes in dystonia and expands the mutational spectrum. Of note, likely pathogenic variants only in GCH1 were also found among PD patients. For DYT-KMT2B, the recently described episignature served as a reliable readout to determine the functional effect of newly identified variants.This work was supported by the German Ministry of Education and Research (BMBF, DYSTRACT consortium, 01GM1514B, to A.A.K., T.B., C.Klein and K.L.) and the German Research Foundation (DFG, LO1555/10-1 to H.B., C.Klein, and K.L. and Project-ID 424778381-TRR 295 to A.A.K). The DysTract registry was further supported by the Arbeitskreis Botulinumtoxin der DGN e.V., Merz Therapeutics, AbbVie/Allergan, and Ipsen Pharma. The Korean DNA samples for this study were provided by the Seoul National University Hospital Human Biobank, a member of the National Biobank of Korea, which is supported by the Ministry of Health and Welfare. All samples derived from the National Biobank of Korea were obtained with informed consent under institutional review board-approved protocols. Several authors are members of the European Reference Network for Rare Neurological Diseases (Project ID No. 739510). Open Access funding enabled and organized by Projekt DEAL.Peer reviewe