Bortezomib in antibody-mediated autoimmune diseases (TAVAB): study protocol for a unicentric, non-randomised, non-placebo controlled trial

Introduction: The clinical characteristics of autoantibodymediated autoimmune diseases are diverse. Yet, medical treatment and the associated complications are similar, that is, the occurrence of long-term side effects and the problem that a significant proportion of patients are non-responders. Therefore, new therapeutic options are needed. Bortezomib, a proteasome inhibitor, is effective in the treatment of multiple myeloma and data from experimental models and case reports suggest an effect in the treatment of autoantibody-mediated autoimmunity. In our study, we will determine the effect of bortezomib treatment on a shared surrogate parameter for clinical efficacy, namely change in autoantibody levels, which we chose as primary parameter. Methods and analysis: We designed a phase IIa trial with altogether n=18 treatment-refractory patients suffering from myasthenia gravis, systemic lupus erythematosus and rheumatoid arthritis that will be treated with bortezomib add-on to pre-existing therapy. Primary endpoint is the change in autoantibody levels 6 months after therapy. Secondary endpoints include concomitant medication, disease-specific clinical scores and measures of quality of life and activities of daily living. Ethics and dissemination: Safety parameters include neurophysiological and clinical signs of peripheral neuropathy as well as potential central nervous system side effects determined by olfactory and neuropsychological testing. The study has been approved by the local ethical committee and first participants have already been enrolled. This proof of concept study will contribute to improve our understanding of plasma cellspecific treatment approaches by assessing its safety and efficacy in reducing serum levels of antibodies known to mediate autoimmune disorders. We plan to publish the final results of our study in a peer reviewed journal and to present our findings at international conferences. Trial registration number: NCT02102594

Effects of spermidine supplementation on cognition and biomarkers in older adults with subjective cognitive decline (SmartAge)—study protocol for a randomized controlled trial

Background: Given the global increase in the aging population and age-related diseases, the promotion of healthy aging is one of the most crucial public health issues. This trial aims to contribute to the establishment of effective approaches to promote cognitive and brain health in older individuals with subjective cognitive decline (SCD). Presence of SCD is known to increase the risk of objective cognitive decline and progression to dementia due to Alzheimer’s disease. Therefore, it is our primary goal to determine whether spermidine supplementation has a positive impact on memory performance in this at-risk group, as compared with placebo. The secondary goal is to examine the effects of spermidine intake on other neuropsychological, behavioral, and physiological parameters. Methods: The SmartAge trial is a monocentric, randomized, double-blind, placebo-controlled phase IIb trial. The study will investigate 12 months of intervention with spermidine-based nutritional supplementation (target intervention) compared with 12months of placebo intake (control intervention). We plan to recruit 100 cognitively normal older individuals with SCD from memory clinics, neurologists and general practitioners in private practice, and the general population. Participants will be allocated to one of the two study arms using blockwise randomization stratified by age and sex with a 1:1 allocation ratio. The primary outcome is the change in memory performance between baseline and post-intervention visits (12 months after baseline). Secondary outcomes include the change in memory performance from baseline to follow-up assessment (18months after baseline), as well as changes in neurocognitive, behavioral, and physiological parameters (including blood and neuroimaging biomarkers), assessed at baseline and post-intervention. Discussion: The SmartAge trial aims to provide evidence of the impact of spermidine supplementation on memory performance in older individuals with SCD. In addition, we will identify possible neurophysiological mechanisms of action underlying the anticipated cognitive benefits. Overall, this trial will contribute to the establishment of nutrition intervention in the prevention of Alzheimer’s disease

Tracking the timely dissemination of clinical studies. Characteristics and impact of 10 tracking variables [version 1; peer review: 2 approved]

Background: Several meta-research studies and benchmarking activities have assessed how comprehensively and timely, academic institutions and private companies publish their clinical studies. These current “clinical trial tracking” activities differ substantially in how they sample relevant studies, and how they follow up on their publication. Methods: To allow informed policy and decision making on future publication assessment and benchmarking of institutions and companies, this paper outlines and discusses 10 variables that influence the tracking of timely publications. Tracking variables were initially selected by experts and by the authors through discussion. To validate the completeness of our set of variables, we conducted i) an explorative review of tracking studies and ii) an explorative tracking of registered clinical trials of three leading German university medical centres. Results: We identified the following 10 relevant variables impacting the tracking of clinical studies: 1) responsibility for clinical studies, 2) type and characteristics of clinical studies, 3) status of clinical studies, 4) source for sampling, 5) timing of registration, 6) determination of completion date, 7) timeliness of dissemination, 8) format of dissemination, 9) source for tracking, and 10) inter-rater reliability. Based on the description of these tracking variables and their influence, we discuss which variables could serve in what ways as a standard assessment of “timely publication”. Conclusions: To facilitate the tracking and consequent benchmarking of how often and how timely academic institutions and private companies publish clinical study results, we have two core recommendations. First, the improvement in the link between registration and publication, for example via institutional policies for academic institutions and private companies. Second, the comprehensive and transparent reporting of tracking studies according to the 10 variables presented in this paper

Effects of Spermidine Supplementation on Cognition and Biomarkers in Older Adults With Subjective Cognitive Decline : Decline A Randomized Clinical Trial

IMPORTANCE Developing interventions against age-related memory decline and for older adults experiencing neurodegenerative disease is one of the greatest challenges of our generation. Spermidine supplementation has shown beneficial effects on brain and cognitive health in animal models, and there has been preliminary evidence of memory improvement in individuals with subjective cognitive decline. OBJECTIVE To determine the effect of longer-term spermidine supplementation on memory performance and biomarkers in this at-risk group. DESIGN, SETTING, AND PARTICIPANTS This 12-month randomized, double-masked, placebocontrolled phase 2b trial (the SmartAge trial) was conducted between January 2017 and May 2020. The study was a monocenter trial carried out at an academic clinical research center in Germany. Eligible individuals were aged 60 to 90 years with subjective cognitive decline who were recruited from health care facilities as well as through advertisements in the general population. Data analysis was conducted between January and March 2021. INTERVENTIONS One hundred participants were randomly assigned (1:1 ratio) to 12 months of dietary supplementation with either a spermidine-rich dietary supplement extracted from wheat germ (O.9 mg spermidine/d) or placebo (microcrystalline cellulose). Eighty-nine participants (89%) successfully completed the trial intervention. MAIN OUTCOMES AND MEASURES Primary outcome was change in memory performance from baseline to 12-month postintervention assessment (intention-to-treat analysis), operationalized by mnemonic discrimination performance assessed by the Mnemonic Similarity Task. Secondary outcomes included additional neuropsychological, behavioral, and physiological parameters. Safety was assessed in all participants and exploratory per-protocol, as well as subgroup, analyses were performed. RESULTS A total of 100 participants (51 in the spermidine group and 49 in the placebo group) were included in the analysis (mean [SD] age, 69 [5] years; 49 female participants [49%]). Over 12 months, no significant changes were observed in mnemonic discrimination performance (between-group difference, -0.03; 95% CI, -0.11 to 0.05; P = .47) and secondary outcomes. Exploratory analyses indicated possible beneficial effects of the intervention on inflammation and verbal memory. Adverse events were balanced between groups. CONCLUSIONS AND RELEVANCE In this randomized clinical trial, longer-term spermidine supplementation in participants with subjective cognitive decline did not modify memory and biomarkers compared with placebo. Exploratory analyses indicated possible beneficial effects on verbal memory and inflammation that need to be validated in future studies at higher dosage.Peer reviewe

Simvastatin add-on to escitalopram in patients with comorbid obesity and major depression (SIMCODE): study protocol of a multicentre, randomised, double-blind, placebo-controlled trial

Introduction: Major depressive disorder (MDD) and obesity are both common disorders associated with significant burden of disease worldwide. Importantly, MDD and obesity often co-occur, with each disorder increasing the risk for developing the other by about 50%-60%. Statins are among the most prescribed medications with well-established safety and efficacy. Statins are recommended in primary prevention of cardiovascular disease, which has been linked to both MDD and obesity. Moreover, statins are promising candidates to treat MDD because a meta-analysis of pilot randomised controlled trials has found antidepressive effects of statins as adjunct therapy to antidepressants. However, no study so far has tested the antidepressive potential of statins in patients with MDD and comorbid obesity. Importantly, this is a difficult-to-treat population that often exhibits a chronic course of MDD and is more likely to be treatment resistant. Thus, in this confirmatory randomised controlled trial, we will determine whether add-on simvastatin to standard antidepressant medication with escitalopram is more efficacious than add-on placebo over 12 weeks in 160 patients with MDD and comorbid obesity. Methods and analysis: This is a protocol for a randomised, placebo-controlled, double-blind multicentre trial with parallel-group design (phase II). One hundred and sixty patients with MDD and comorbid obesity will be randomised 1:1 to simvastatin or placebo as add-on to standard antidepressant medication with escitalopram. The primary outcome is change in the Montgomery-angstrom sberg Depression Rating Scale (MADRS) score from baseline to week 12. Secondary outcomes include MADRS response (defined as 50% MADRS score reduction from baseline), MADRS remission (defined as MADRS score <10), mean change in patients' self-reported Beck Depression Inventory (BDI-II) and mean change in high-density lipoprotein, low-density lipoprotein and total cholesterol from baseline to week 12. Ethics and dissemination: This protocol has been approved by the ethics committee of the federal state of Berlin (Ethik-Kommission des Landes Berlin, reference: 19/0226-EK 11) and by the relevant federal authority (Bundesinstitut fur Arzneimittel und Medizinprodukte (BfArM), reference: 4043387). Study findings will be published in peer-reviewed journals and will be presented at (inter)national conferences

Rationale and design of the prevention of paclitaxel-related neurological side effects with lithium trial – Protocol of a multicenter, randomized, double-blind, placebo-controlled proof-of-concept phase-2 clinical trial

INTRODUCTION: Chemotherapy-induced polyneuropathy (CIPN) and post-chemotherapy cognitive impairment (PCCI) are frequent side effects of paclitaxel treatment. CIPN/PCCI are potentially irreversible, reduce quality of life and often lead to treatment limitations, which affect patients’ outcome. We previously demonstrated that paclitaxel enhances an interaction of the Neuronal calcium sensor-1 protein (NCS-1) with the Inositol-1,4,5-trisphosphate receptor (InsP3R), which disrupts calcium homeostasis and triggers neuronal cell death via the calcium-dependent protease calpain in dorsal root ganglia neurons and neuronal precursor cells. Prophylactic treatment of rodents with lithium inhibits the NCS1-InsP3R interaction and ameliorates paclitaxel-induced polyneuropathy and cognitive impairment, which is in part supported by limited retrospective clinical data in patients treated with lithium carbonate at the time of chemotherapy. Currently no data are available from a prospective clinical trial to demonstrate its efficacy. METHODS AND ANALYSIS: The PREPARE study will be conducted as a multicenter, randomized, double-blind, placebo-controlled phase-2 trial with parallel group design. N = 84 patients with breast cancer will be randomized 1:1 to either lithium carbonate treatment (targeted serum concentration 0.5–0.8 mmol/l) or placebo with sham dose adjustments as add-on to (nab-) paclitaxel. The primary endpoint is the validated Total Neuropathy Score reduced (TNSr) at 2 weeks after the last (nab-) paclitaxel infusion. The aim is to show that the lithium carbonate group is superior to the placebo group, meaning that the mean TNSr after (nab-) paclitaxel is lower in the lithium carbonate group than in the placebo group. Secondary endpoints include: (1) severity of CIPN, (2) amount and dose of pain medication, (3) cumulative dose of (nab-) paclitaxel, (4) patient-reported symptoms of CIPN, quality of life and symptoms of anxiety and depression, (5) severity of cognitive impairment, (6) hippocampal volume and changes in structural/functional connectivity and (7) serum Neurofilament light chain protein concentrations. ETHICS AND DISSEMINATION: The study protocol was approved by the Berlin ethics committee (reference: 21/232 – IV E 10) and the respective federal agency (Bundesinstitut für Arzneimittel und Medizinprodukte, reference: 61-3910-4044771). The results of the study will be published in peer-reviewed medical journals as well as presented at relevant (inter)national conferences. CLINICAL TRIAL REGISTRATION: [https://www.drks.de/drks_web/navigate.do?navigationId=trial.HTML&TRIAL_ID=DRKS00027165], identifier [DRKS00027165]

Funktionelle Analyse von komplexen Hepatitis-B-Virus-Varianten, assoziiert mit Leberzirrhose bei Immunsupprimierten

Obwohl der Wildtyp des Hepatitis-B-Virus (HBV) nicht zytopathogen und die Pathogenese der Hepatitis B generell immunvermittelt ist, können in immunsupprimierten Nierentransplantatempfängern mit chronischer Hepatitis B schwere Leberschäden bis hin zu Leberzirrhose und Leberversagen entstehen. Die Entwicklung von Leberzirrhose in den Nierentransplantierten ist assoziiert mit der Akkumulation und Persistenz von komplexen HBV-Varianten mit Mutationen im Core-Promotor / X-Gen, Deletionen im Core (C)-Gen und teilweise zusätzlichen Deletionen im präS-Bereich. Dies lässt eine Rolle der Varianten in der speziellen Pathogenese bei Immunsupprimierten vermuten. In der vorliegenden Arbeit wurden funktionelle Analysen der komplexen Varianten im Vergleich zu Referenz-Wildtypgenomen und Wildtyp-ähnlichen Genomen der Patienten aus der frühen Infektionsphase durchgeführt, um Hinweise auf den potentiellen Beitrag der Varianten zur Pathogenese zu erlangen. Die Analysen erfolgten durch transiente Transfektion der humanen Hepatomazelllinie HuH7 mit repräsentativen HBV-Gesamtgenomen, die aus 2 Patienten während des Krankheitsverlaufs von einer asymptomatischen Infektion hin zur Leberzirrhose isoliert und kloniert worden waren. Trotz einiger Unterschiede im Detail wiesen die komplexen Varianten einen gemeinsamen, drastisch vom Wildtyp abweichenden Phänotyp auf. Dieser war gekennzeichnet durch eine veränderte Transkription mit reduzierten präC- und Oberflächen-mRNAs und verstärkter Expression der prägenomischen RNA, eine starke Reduktion des häufigsten Spleißprodukts der prägenomischen RNA, SP1, eine extrem reduzierte oder fehlende Expression und/oder Sekretion aller Oberflächenproteine und des HBeAg, ein verändertes intrazelluläres Verteilungsmuster des schwach exprimierten Core-Proteins und teilweise der Oberflächenproteine sowie eine erhöhte Replikation und Anreicherung gegenüber Wildtyp-HBV aufgrund einer verstärkten reversen Transkription der prägenomischen RNA. Dieser Phänotyp basierte zum Teil auf den Mutationen in Core-Promotor und C-Gen, wurde jedoch deutlich durch zusätzliche Mutationen in den übrigen Genomabschnitten beeinflusst. Die vielfältigen Veränderungen der Varianten unterstützen ihren vermuteten Beitrag zur Pathogenese.Although wild-type hepatitis B virus is not cytopathogenic and the pathogenesis of hepatitis B is generally immune mediated, also immuno-suppressed patients, such as renal transplant recipients, with chronic hepatitis B may develop liver cirrhosis and end-stage liver disease. In renal transplant recipients, the development of liver cirrhosis is associated with the accumulation and persistence of complex HBV variants with mutations in core promoter / X gene, deletions in core (C) gene and sometimes additional deletions in the preS region. This suggests a role of these variants in the special pathogenesis in immuno-suppressed patients. In the present work, the complex variants were functionally analyzed in comparison to reference wild-type genomes and wild-type-like HBV genomes from the early asymptomatic phase of infection. For the analyses, representative cloned full-length HBV genomes isolated from 2 patients before and during liver cirrhosis were transiently transfected into the human hepatoma cell line HuH7. In spite of some variations, the complex variants showed a common phenotype, which was drastically altered compared to wild-type. It was characterized by reduced preC and surface mRNAs and increased expression of pregenomic RNA, by a strong reduction of the major spliced pregenomic RNA, SP1, by a partial or complete defect in expression and/or secretion of surface proteins and HBeAg, by an aberrant intracellular localization of the weakly expressed core protein and in some cases of the surface proteins, and by an enhanced replication and enrichment over wild-type HBV due to an enhanced reverse transcription of variant pregenomic RNA. The phenotypic alterations were often based on the mutations in core promoter and C gene but were considerably influenced by the additional mutations in other genomic regions. The multiple functional changes of the variants support their assumed contribution to pathogenesis

New purines with activity of coagulating

Titelblatt Inhaltsverzeichnis Einleitung 1 Chemisch-theoretischer Teil 7 Pharmakologischer Teil 57 Chemisch-experimenteller Teil 89 Zusammenfassung 149 Literaturverzeichnis 155 Verzeichnis der beschriebenen Substanzen 161Ziel dieser Arbeit war die Synthese von neuen Substanzen mit Purin- Grundgerüst, die über eine Erhöhung des cGMP-Spiegels (cyclisches Guanosinmonophosphat) die Thrombozyten-aggregation hemmen. Nach Vergleich von Aktivatoren der löslichen Guanylatcyclase (sGC) wie YC-1 und Inhibitoren der Phosphodiesterase vom Typ 5 (PDE 5) wie Zaprinast und Sildenafil konnte aufgrund von strukturellen Ahnlichkeiten ein Arbeitsschema entwickelt werden, um Substanzen darzustellen, die beide Enzyme beeinflussen. So sollte eine synergistische, cGMP-erhöhende Wirkung erzielt werden. Nach diesem Arbeitsschema wurden zuerst Purin-2,6-diamine mit basischen (6-10), neutralen (11-13) oder sauren (14) Substituenten in der N6-Alkylseitenkette dargestellt. Im Born-Test, einem in-vitro-Testmodell zur Bestimmung der antiaggregatorischen Aktivität, zeigten vorwiegend die Substanzen mit basischem Rest in der Seitenkette unter Zusatz von Kollagen als Aggregationsauslöser thrombozytenaggregationshemmende Eigenschaften (IC50 um 200 µmol/L). Die Umsetzung zu den Purin-2-onen (16-18) bzw. N-(Purin-2-yl)benzolsulfonamiden (20,21) führte fast ausschließlich zu höheren IC50-Werten und damit zum Wirkungsverlust. Im zweiten Teil der Arbeit wurden diverse N-(Purin-2-yl)benzolcarbonsäureamide mit (25,26) und ohne (27,28) weiterer Sulfonamid-Partialstruktur dargestellt. Dies führte zu einer deutlichen Steigerung der antiaggregatorischen Aktivität. Hier war die Substanz 25h mit einer IC50 von 43 µmol/L am aktivsten. Um stärker auf die Löslichkeitseigenschaften der untersuchten Substanzen einzugehen, wurde im Born-Test die Inkubationszeit auf 20 min erhöht. Dies führte bei 25h zu einer Senkung des IC50-Wertes auf 10 µmol/L. Eine Auswahl von in vitro effektiven Substanzen wurden auf die Hemmung der PDE 5 und die Aktivierung der sGC untersucht. Dabei stellte sich heraus, daß die beobachteten Hemmeffekte nicht auf eine Beeinflussung dieser beiden Enzyme zurückzuführen war. Bei der Zugabe von PAF (Plättchenaktivierender Faktor) als Aggregationsauslöser im Born-Test wurde die IC50 bei 25h von 10 µmol/L auf 1.4 µmol/L gesenkt. Damit konnte die Hemmung der Thrombozytenaggregation von 25h auf einen PAF-Rezeptor- Antagonismus zurückgeführt werden. Mit Hilfe eines Laser-Thrombose-Modells wurden ausgewählte Substanzen in vivo auf die Hemmung der Thrombusbildung in den Arteriolen und Venolen von Ratten untersucht. Hier war 21 die aktivste Substanz. Sie zeigte 2h nach peroraler Gabe (60 mg/kg) eine Hemmung auf die Thrombusbildung von 8 % in den Arteriolen und 4 % in den Venolen.Aim of this thesis was the synthesis of new substances with Purine basic body that inhibit the platelet aggregation by an increase in concentration of cGMP (cyclic Guanosinmonophosphate). After comparing activators of the soluble guanylat cyclase (sGC) like YC-1 and inhibitors of the phosphodiesterase of type 5 (PDE 5) like Zaprinast and Sildenafil a work model based on structural similarities was developed to synthesize substances that influence both enzymes. This promises a synergistic, cGMP-raising, effect. According to this work model Purin-2,6-diamine with basic (6-10), neutral (11-13) or acid (14) group at the alkyl side chain was prepared. For the determination of platelet aggregation in vitro the Born-Test was used. At the Born-Test chiefly compounds with basic group were shown to inhibit the collagen-induced platelet aggregation (IC50 ≈ 200 µmol/L). The synthesis of Purin-2-one (16-18) and N-(Purin-2-yl)-benzolsulfonamide (20,21) led to a loss of activity (IC50 > 300 µmol/L). At the second part of this thesis N-(Purin-2-yl)-benzolcarboxamide with (25,26) and without (27,28) Sulfonamide-partialstructures were prepared. These compounds led to an increase of inhibition of platelet aggregation at the Born-Test. Substance 25h was the most active benzolcarboxamide with collagen as inductor, that was obtained (IC50 = 43 µmol/L). Prolongation of incubation time from 4 minutes to 20 minutes led to increase in activity (i.e. 25h:IC50 = 10 µmol/L). A selection of in vitro effective substances was also tested for inhibition of PDE 5 and activation of sGC. None of these compounds showed inhibition of PDE 5 or activation of sGC. When platelet activating factor (PAF) was used to induce the platelet aggregation at the Born-Test the IC50 from substance 25h decreased from 10 µmol/L (collagen) to 1.4 µmol/L. This showed, that 25h is a PAF-receptor-antagonist. By use of the Laser- Thrombosis-Model some selected compounds were tested for antithrombotic effects in vivo. The most active substance was 21. 2 hour after peroral application (60 mg/kg) it reduced the formation of thrombi in arterioles by 8 % and venoles by 4 %