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Grau, Janina ; Zöllner, Johann Philipp ; Schubert-Bast, Susanne ; Kurlemann, Gerhard ; Hertzberg, Christoph ; Wiemer-Kruel, Adelheid ; Bast, Thomas ; Bettendorf, Ulrich ; Fiedler, Barbara ; Hahn, Andreas ; Hartmann, Hans ; Hornemann, Frauke ; Immisch, Ilka ; Jacobs, Julia ; Kieslich, Matthias ; Klein, Karl Martin ; Klotz, Kerstin Alexandra ; Kluger, Gerhard ; Knuf, Markus ; Mayer, Thomas ; Marquard, Klaus ; Meyer, Sascha ; Muhle, Hiltrud ; Müller-Schlüter, Karen ; Noda, Anna ; Ruf, Susanne ; Sauter, Matthias ; Schlump, Jan-Ulrich ; Syrbe, Steffen ; Thiels, Charlotte ; Trollmann, Regina ; Wilken, Bernd ; Willems, Laurent Maximilian ; Rosenow, Felix ; Strzelczyk, Adam
Background: Tuberous sclerosis complex (TSC), a multisystem genetic disorder, affects many organs and systems, characterized by benign growths. This German multicenter study estimated the disease-specific costs and cost-driving factors associated with various organ manifestations in TSC patients. Methods: A validated, three-month, retrospective questionnaire was administered to assess the sociodemographic and clinical characteristics, organ manifestations, direct, indirect, out-of-pocket, and nursing care-level costs, completed by caregivers of patients with TSC throughout Germany. Results: The caregivers of 184 patients (mean age 9.8 ± 5.3 years, range 0.7–21.8 years) submitted questionnaires. The reported TSC disease manifestations included epilepsy (92%), skin disorders (86%), structural brain disorders (83%), heart and circulatory system disorders (67%), kidney and urinary tract disorders (53%), and psychiatric disorders (51%). Genetic variations in TSC2 were reported in 46% of patients, whereas 14% were reported in TSC1. Mean total direct health care costs were EUR 4949 [95% confidence interval (95% CI) EUR 4088–5863, median EUR 2062] per patient over three months. Medication costs represented the largest direct cost category (54% of total direct costs, mean EUR 2658), with mechanistic target of rapamycin (mTOR) inhibitors representing the largest share (47%, EUR 2309). The cost of anti-seizure drugs (ASDs) accounted for a mean of only EUR 260 (5%). Inpatient costs (21%, EUR 1027) and ancillary therapy costs (8%, EUR 407) were also important direct cost components. The mean nursing care-level costs were EUR 1163 (95% CI EUR 1027–1314, median EUR 1635) over three months. Total indirect costs totaled a mean of EUR 2813 (95% CI EUR 2221–3394, median EUR 215) for mothers and EUR 372 (95% CI EUR 193–586, median EUR 0) for fathers. Multiple regression analyses revealed polytherapy with two or more ASDs and the use of mTOR inhibitors as independent cost-driving factors of total direct costs. Disability and psychiatric disease were independent cost-driving factors for total indirect costs as well as for nursing care-level costs. Conclusions: This study revealed substantial direct (including medication), nursing care-level, and indirect costs associated with TSC over three months, highlighting the spectrum of organ manifestations and their treatment needs in the German healthcare setting.
Herzer, Kerstin ; Sterneck, Martina ; Welker, Martin-Walter ; Nadalin, Silvio ; Kirchner, Gabriele ; Braun, Felix ; Malessa, Christina ; Herber, Adam ; Pratschke, Johann ; Weiss, Karl Heinz ; Jäckel, Elmar ; Tacke, Frank
Improving long-term patient and graft survival after liver transplantation (LT) remains a major challenge. Compared to the early phase after LT, long-term morbidity and mortality of the recipients not only depends on complications immediately related to the graft function, infections, or rejection, but also on medical factors such as de novo malignancies, metabolic disorders (e.g., new-onset diabetes, osteoporosis), psychiatric conditions (e.g., anxiety, depression), renal failure, and cardiovascular diseases. While a comprehensive post-transplant care at the LT center and the connected regional networks may improve outcome, there is currently no generally accepted standard to the post-transplant management of LT recipients in Germany. We therefore described the structure and standards of post-LT care by conducting a survey at 12 German LT centers including transplant hepatologists and surgeons. Aftercare structures and form of cost reimbursement considerably varied between LT centers across Germany. Further discussions and studies are required to define optimal structure and content of post-LT care systems, aiming at improving the long-term outcomes of LT recipients.
Wurm, Jan Philip ; Meyer, Britta ; Bahr, Ute ; Held, Martin ; Frolow, Olga ; Kötter, Peter ; Engels, Joachim W. ; Heckel, Alexander ; Karas, Michael ; Entian, Karl-Dieter ; Wöhnert, Jens
Nep1 (Emg1) is a highly conserved nucleolar protein with an essential function in ribosome biogenesis. A mutation in the human Nep1 homolog causes Bowen–Conradi syndrome—a severe developmental disorder. Structures of Nep1 revealed a dimer with a fold similar to the SPOUT-class of RNA-methyltransferases suggesting that Nep1 acts as a methyltransferase in ribosome biogenesis. The target for this putative methyltransferase activity has not been identified yet. We characterized the RNA-binding specificity of Methanocaldococcus jannaschii Nep1 by fluorescence- and NMR-spectroscopy as well as by yeast three-hybrid screening. Nep1 binds with high affinity to short RNA oligonucleotides corresponding to nt 910–921 of M. jannaschii 16S rRNA through a highly conserved basic surface cleft along the dimer interface. Nep1 only methylates RNAs containing a pseudouridine at a position corresponding to a previously identified hypermodified N1-methyl-N3-(3-amino-3-carboxypropyl) pseudouridine (m1acp3-Psi) in eukaryotic 18S rRNAs. Analysis of the methylated nucleoside by MALDI-mass spectrometry, HPLC and NMR shows that the methyl group is transferred to the N1 of the pseudouridine. Thus, Nep1 is the first identified example of an N1-specific pseudouridine methyltransferase. This enzymatic activity is also conserved in human Nep1 suggesting that Nep1 is the methyltransferase in the biosynthesis of m1acp3-Psi in eukaryotic 18S rRNAs.
Miethke, Marcus ; Pieroni, Marco ; Weber, Karl Tilmann ; Brönstrup, Mark ; Hammann, Peter ; Halby, Ludovic ; Arimondo, Paola B. ; Glaser, Philippe ; Aigle, Bertrand ; Bode, Helge Björn ; Moreira, Rui ; Li, Yanyan ; Luzhetskyy, Andriy ; Medema, Marnix H. ; Pernodet, Jean-Luc ; Stadler, Marc ; Tormo, José Rubén ; Genilloud, Olga ; Truman, Andrew W. ; Weissman, Kira J. ; Takano, Eriko ; Sabatini, Stefano ; Stegmann, Evi ; Brötz, Heike ; Wohlleben, Wolfgang ; Seemann, Myriam ; Empting, Martin ; Hirsch, Anna K. H. ; Loretz, Brigitta ; Lehr, Claus-Michael ; Titz, Alexander ; Herrmann, Jennifer ; Jäger, Timo ; Alt, Silke ; Hesterkamp, Thomas ; Winterhalter, Mathias ; Schiefer, Andrea ; Pfarr, Kenneth ; Hoerauf, Achim ; Graz, Heather ; Graz, Michael ; Lindvall, Mika ; Ramurthy, Savithri ; Karlén, Anders ; Dongen, Maarten van ; Petkovic, Hrvoje ; Keller, Andreas ; Peyrane, Frédéric ; Donadio, Stefano ; Fraisse, Laurent ; Piddock, Laura J. V. ; Gilbert, Ian H. ; Moser, Heinz E. ; Müller, Rolf
An ever-increasing demand for novel antimicrobials to treat life-threatening infections caused by the global spread of multidrug-resistant bacterial pathogens stands in stark contrast to the current level of investment in their development, particularly in the fields of natural-product-derived and synthetic small molecules. New agents displaying innovative chemistry and modes of action are desperately needed worldwide to tackle the public health menace posed by antimicrobial resistance. Here, our consortium presents a strategic blueprint to substantially improve our ability to discover and develop new antibiotics. We propose both short-term and long-term solutions to overcome the most urgent limitations in the various sectors of research and funding, aiming to bridge the gap between academic, industrial and political stakeholders, and to unite interdisciplinary expertise in order to efficiently fuel the translational pipeline for the benefit of future generations.
Meyer, Britta ; Wurm, Jan Philip ; Kötter, Peter ; Leisegang, Matthias ; Schilling, Valeska ; Buchhaupt, Markus ; Held, Martin ; Bahr, Ute ; Karas, Michael ; Heckel, Alexander ; Bohnsack, Markus ; Wöhnert, Jens ; Entian, Karl-Dieter
The Nep1 (Emg1) SPOUT-class methyltransferase is an essential ribosome assembly factor and the human Bowen–Conradi syndrome (BCS) is caused by a specific Nep1D86G mutation. We recently showed in vitro that Methanocaldococcus jannaschii Nep1 is a sequence-specific pseudouridine-N1-methyltransferase. Here, we show that in yeast the in vivo target site for Nep1-catalyzed methylation is located within loop 35 of the 18S rRNA that contains the unique hypermodification of U1191 to 1-methyl-3-(3-amino-3-carboxypropyl)-pseudouri-dine (m1acp3Psi). Specific 14C-methionine labelling of 18S rRNA in yeast mutants showed that Nep1 is not required for acp-modification but suggested a function in Psi1191 methylation. ESI MS analysis of acp-modified Psi-nucleosides in a DeltaNep1-mutant showed that Nep1 catalyzes the Psi1191 methylation in vivo. Remarkably, the restored growth of a nep1-1ts mutant upon addition of S-adenosylmethionine was even observed after preventing U1191 methylation in a deltasnr35 mutant. This strongly suggests a dual Nep1 function, as Psi1191-methyltransferase and ribosome assembly factor. Interestingly, the Nep1 methyltransferase activity is not affected upon introduction of the BCS mutation. Instead, the mutated protein shows enhanced dimerization propensity and increased affinity for its RNA-target in vitro. Furthermore, the BCS mutation prevents nucleolar accumulation of Nep1, which could be the reason for reduced growth in yeast and the Bowen-Conradi syndrome.
Pratschke, Sebastian ; Eder, Michael ; Heise, Michael ; Nadalin, Silvio ; Pascher, Andreas ; Schemmer, Peter ; Scherer, Marcus N. ; Ulrich, Frank ; Wolters, Heiner ; Jauch, Karl-Walter ; Wöhling, Dirk ; Angele, Martin Kurt
Background: Critical organ shortage results in the utilization of extended donor criteria (EDC) liver grafts. These marginal liver grafts are prone to increased ischemia reperfusion injury (IRI) which may contribute to deteriorated graft function and survival. Experimental data have shown that the calcineurin inhibitor tacrolimus exerts protective effects on hepatic IRI when applied intravenously or directly as a hepatic rinse. Therefore, the aim of the present study is to examine the effects of an ex vivo tacrolimus perfusion on IRI in transplantation of EDC liver grafts.
Methods/Design: The TOP-Study (tacrolimus organ perfusion) is a randomized multicenter trial comparing the ex vivo tacrolimus perfusion of marginal liver grafts with placebo. We hypothesize that a tacrolimus rinse reduces IRI, potentially improving organ survival following transplantation of EDC livers. The study includes livers with two or more EDC, according to Eurotransplant International Foundation’s definition of EDC livers. Prior to implantation, livers randomized to the treatment group are rinsed with tacrolimus at a concentration of 20 ng/ml in 1000 ml Custodiol solution and in the placebo group with Custodiol alone. The primary endpoint is the maximum serum alanine transamninase (ALT) level within the first 48 hours after surgery; however, the study design also includes a 1-year observation period following transplantation. The TOP-Study is an investigator-initiated trial sponsored by the University of Munich Hospital. Seven other German transplant centers are participating (Berlin, Frankfurt, Heidelberg, Mainz, Münster, Regensburg, Tübingen) and aim to include a total of 86 patients.
Discussion: Tacrolimus organ perfusion represents a promising strategy to reduce hepatic IRI following the transplantation of marginal liver grafts. This treatment may help to improve the function of EDC grafts and therefore safely expand the donor pool in light of critical organ shortage.
Trial register: EudraCT number: 2010-021333-31, ClinicalTrials.gov identifier: NCT01564095
Bönig, Halvard-Björn ; Kuçi, Zyrafete ; Kuçi, Selim ; Bakhtiar, Shahrzad ; Basu, Oliver ; Bug, Gesine ; Dennis, Mike ; Greil, Johann ; Barta, Aniko ; Kállay, Krisztián ; Lang, Peter ; Lucchini, Giovanna ; Pol, Raj ; Schulz, Ansgar ; Sykora, Karl-Walter ; Teichert- von Lüttichau, Irene ; Herter-Sprie, Grit ; Uddin, Mohammad Ashab ; Jenkin, Phil ; Alsultan, Abdulrahman ; Büchner, Jochen ; Stein, Jerry ; Kelemen, Agnes ; Jarisch, Andrea ; Sörensen, Jan ; Salzmann-Manrique, Emilia ; Hutter, Martin ; Schäfer, Richard ; Seifried, Erhard ; Paneesha, Shankara ; Novitzky-Basso, Igor ; Gefen, Aharon ; Nevo, Neta ; Beutel, Gernot ; Schlegel, Paul-Gerhardt ; Klingebiel, Thomas ; Bader, Peter
(1) Background: Refractory acute graft-versus-host disease (R-aGvHD) remains a leading cause of death after allogeneic stem cell transplantation. Survival rates of 15% after four years are currently achieved; deaths are only in part due to aGvHD itself, but mostly due to adverse effects of R-aGvHD treatment with immunosuppressive agents as these predispose patients to opportunistic infections and loss of graft-versus-leukemia surveillance resulting in relapse. Mesenchymal stromal cells (MSC) from different tissues and those generated by various protocols have been proposed as a remedy for R-aGvHD but the enthusiasm raised by initial reports has not been ubiquitously reproduced.
(2) Methods: We previously reported on a unique MSC product, which was generated from pooled bone marrow mononuclear cells of multiple third-party donors. The products showed dose-to-dose equipotency and greater immunosuppressive capacity than individually expanded MSCs from the same donors. This product, MSC-FFM, has entered clinical routine in Germany where it is licensed with a national hospital exemption authorization. We previously reported satisfying initial clinical outcomes, which we are now updating. The data were collected in our post-approval pharmacovigilance program, i.e., this is not a clinical study and the data is high-level and non-monitored.
(3) Results: Follow-up for 92 recipients of MSC-FFM was reported, 88 with GvHD ≥°III, one-third only steroid-refractory and two-thirds therapy resistant (refractory to steroids plus ≥2 additional lines of treatment). A median of three doses of MSC-FFM was administered without apparent toxicity. Overall response rates were 82% and 81% at the first and last evaluation, respectively. At six months, the estimated overall survival was 64%, while the cumulative incidence of death from underlying disease was 3%.
(4) Conclusions: MSC-FFM promises to be a safe and efficient treatment for severe R-aGvHD.
Kron, Anna ; Alidousty, Christina ; Scheffler, Matthias ; Merkelbach-Bruse, Sabine ; Seidel, Danila ; Riedel, Richard ; Ihle, Michaela A. ; Michels, Sebastian Yves Friedrich ; Nogova, Lucia ; Fassunke, Jana ; Heydt, Carina ; Kron, Florian ; Ueckeroth, Frank ; Serke, Monika ; Krüger, Stefan ; Grohé, Christian ; Koschel, Dirk ; Benedikter, Josef ; Kaminsky, Britta ; Schaaf, Bernhard ; Braess, Jan ; Sebastian, Martin ; Kambartel, Karl-Otto ; Thomas, Roman ; Zander, Thomas ; Schultheis, Anne Maria ; Büttner, Reinhard ; Wolf, Jürgen
Background: We analyzed whether co-occurring mutations influence the outcome of systemic therapy in ALK-rearranged non-small-cell lung cancer (NSCLC).
Patients and methods: ALK-rearranged stage IIIB/IV NSCLC patients were analyzed with next-generation sequencing and fluorescence in situ hybridization analyses on a centralized diagnostic platform. Median progression-free survival (PFS) and overall survival (OS) were determined in the total cohort and in treatment-related sub-cohorts. Cox regression analyses were carried out to exclude confounders.
Results: Among 216 patients with ALK-rearranged NSCLC, the frequency of pathogenic TP53 mutations was 23.8%, while other co-occurring mutations were rare events. In ALK/TP53 co-mutated patients, median PFS and OS were significantly lower compared with TP53 wildtype patients [PFS 3.9 months (95% CI: 2.4–5.6) versus 10.3 months (95% CI: 8.6–12.0), P < 0.001; OS 15.0 months (95% CI: 5.0–24.9) versus 50.0 months (95% CI: 22.9–77.1), P = 0.002]. This difference was confirmed in all treatment-related subgroups including chemotherapy only [PFS first-line chemotherapy 2.6 months (95% CI: 1.3–4.1) versus 6.2 months (95% CI: 1.8–10.5), P = 0.021; OS 2.0 months (95% CI: 0.0–4.6) versus 9.0 months (95% CI: 6.1–11.9), P = 0.035], crizotinib plus chemotherapy [PFS crizotinib 5.0 months (95% CI: 2.9–7.2) versus 14.0 months (95% CI: 8.0–20.1), P < 0.001; OS 17.0 months (95% CI: 6.7–27.3) versus not reached, P = 0.049] and crizotinib followed by next-generation ALK-inhibitor [PFS next-generation inhibitor 5.4 months (95% CI: 0.1–10.7) versus 9.9 months (95% CI: 6.4–13.5), P = 0.039; OS 7.0 months versus 50.0 months (95% CI: not reached), P = 0.001).
Conclusions: In ALK-rearranged NSCLC co-occurring TP53 mutations predict an unfavorable outcome of systemic therapy. Our observations encourage future research to understand the underlying molecular mechanisms and to improve treatment outcome of the ALK/TP53 co-mutated subgroup.
Benz, Fabienne ; Wichitnaowarat, Viraya ; Lehmann, Martin ; Germano, Raoul F. V. ; Mihova, Diana ; Macas, Jadranka ; Adams, Ralf Heinrich ; Taketo, Makoto Mark ; Guerit, Sylvaine ; Plate, Karl ; Vanhollebeke, Benoit ; Liebner, Stefan
The circumventricular organs (CVOs) in the central nervous system (CNS) lack a vascular blood-brain barrier (BBB), creating communication sites for sensory or secretory neurons, involved in body homeostasis. Wnt/β-catenin signaling is essential for BBB development and maintenance in endothelial cells (ECs) in most CNS vessels. Here we show that in mouse development, as well as in adult mouse and zebrafish, CVO ECs rendered Wnt-reporter negative, suggesting low level pathway activity. Characterization of the subfornical organ (SFO) vasculature revealed heterogenous claudin-5 (Cldn5) and Plvap/Meca32 expression indicative for tight and leaky vessels, respectively. Dominant, EC-specific β-catenin transcription in mice, converted phenotypically leaky into BBB-like vessels, by augmenting Cldn5+ vessels, stabilizing junctions and by reducing Plvap/Meca32+ and fenestrated vessels, resulting in decreased tracer permeability. Endothelial tightening augmented neuronal activity in the SFO of water restricted mice. Hence, regulating the SFO vessel barrier may influence neuronal function in the context of water homeostasis.
Blum, Andrea ; Khalifa, Saleem ; Nordström, Karl ; Simon, Martin ; Schulz, Marcel Holger ; Schmit, Manfred J.
KDEL receptors (KDELRs) represent transmembrane proteins of the secretory pathway which regulate the retention of soluble ER-residents as well as retrograde and anterograde vesicle trafficking. In addition, KDELRs are involved in the regulation of cellular stress response and ECM degradation. For a deeper insight into KDELR1 specific functions, we characterised a KDELR1-KO cell line (HAP1) through whole transcriptome analysis by comparing KDELR1-KO cells with its respective HAP1 wild-type. Our data indicate more than 300 significantly and differentially expressed genes whose gene products are mainly involved in developmental processes such as cell adhesion and ECM composition, pointing out to severe cellular disorders due to a loss of KDELR1. Impaired adhesion capacity of KDELR1-KO cells was further demonstrated through in vitro adhesion assays, while collagen- and/or laminin-coating nearly doubled the adhesion property of KDELR1-KO cells compared to wild-type, confirming a transcriptional adaptation to improve or restore the cellular adhesion capability. Perturbations within the secretory pathway were verified by an increased secretion of ER-resident PDI and decreased cell viability under ER stress conditions, suggesting KDELR1-KO cells to be severely impaired in maintaining cellular homeostasis.