TY - JOUR A1 - Ziegler, Sabrina A1 - Weiss, Esther A1 - Schmitt, Anna-Lena A1 - Schlegel, Jan A1 - Burgert, Anne A1 - Terpitz, Ulrich A1 - Sauer, Markus A1 - Moretta, Lorenzo A1 - Sivori, Simona A1 - Leonhardt, Ines A1 - Kurzai, Oliver A1 - Einsele, Hermann A1 - Loeffler, Juergen T1 - CD56 Is a Pathogen Recognition Receptor on Human Natural Killer Cells JF - Scientific Reports N2 - Aspergillus (A.) fumigatus is an opportunistic fungal mold inducing invasive aspergillosis (IA) in immunocompromised patients. Although antifungal activity of human natural killer (NK) cells was shown in previous studies, the underlying cellular mechanisms and pathogen recognition receptors (PRRs) are still unknown. Using flow cytometry we were able to show that the fluorescence positivity of the surface receptor CD56 significantly decreased upon fungal contact. To visualize the interaction site of NK cells and A. fumigatus we used SEM, CLSM and dSTORM techniques, which clearly demonstrated that NK cells directly interact with A. fumigatus via CD56 and that CD56 is re-organized and accumulated at this interaction site time-dependently. The inhibition of the cytoskeleton showed that the receptor re-organization was an active process dependent on actin re-arrangements. Furthermore, we could show that CD56 plays a role in the fungus mediated NK cell activation, since blocking of CD56 surface receptor reduced fungal mediated NK cell activation and reduced cytokine secretion. These results confirmed the direct interaction of NK cells and A. fumigatus, leading to the conclusion that CD56 is a pathogen recognition receptor. These findings give new insights into the functional role of CD56 in the pathogen recognition during the innate immune response. KW - pattern recognition receptors KW - fungal infection KW - Aspergillus fumigatus KW - natural killer cells Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-170637 VL - 7 IS - 6138 ER - TY - THES A1 - Klein [geb. Geiger], Christina T1 - Entwicklung und Charakterisierung von TNFR2-spezifischen Agonisten T1 - Development and characterisation of TNFR2-specific agonists N2 - Liganden und Rezeptoren des Körpers spielen eine multifaktorielle Rolle in der Regulierung zellulärer Prozesse des Körpers. Der Tumornekrosefaktor (TNF), ein proinflammatorisches Zytokin, bindet natürlicherweise an zwei Rezeptoren, den TNF-Rezeptor 1 (TNFR1) oder den TNFR-Rezeptor 2 (TNFR2) und kann durch Aktivierung vielfältiger Signalwege unterschiedliche Zelleffekte im Körper auslösen. Während TNF in membrangebundener Form vorkommend TNFR1 sowie TNFR2 optimal stimulieren kann, ist lösliches TNF in der Lage zwar an beide Rezeptoren zu binden, natürlicherweise jedoch nur den TNFR1 zu stimulieren. Da eine unkontrollierte Bindung bzw. Aktivierung von beiden Rezeptoren schwere unerwünschte Nebenwirkungen wie Inflammationen haben kann, wurden zur konkreten Aktivierung der einzelnen Rezeptoren TNFR1 und TNFR2 spezifische TNF-Mutanten, wie TNF80 zur Bindung an TNFR2 und TNF60 zur Bindung an TNFR1 konstruiert. Durch die TNF-Mutante TNF80 gelingt es die TNFR2 Wirkungskette zu aktivieren, während die TNFR1-Stimulation verhindert wird. Die Aktivierung des TNFR2-Rezeptors hat eine Stimulierung von regulatorischen T-Zellen (Tregs) zur Folge. Im Rahmen dieser Dissertation wurden einerseits die TNF-TNC-Formen weiterentwickelt, indem die konstante Domäne der schweren Antikörperkette des humanen IgG1 (Fc) hinzukloniert wurde. Hier wurde primär der Effekt der Oligomerisierung mit der aktivierenden Wirkung auf TNFR2 erforscht. Weiterhin wird jedoch durch die Bindungsspezifität des Fc-Fusionsproteins von TNF80 an Tregs eine antitumorale Wirkung ausgelöst, indem durch das ausgelöste ADCC die Tregs zerstört werden. Andererseits wurden Kombinationskonstrukte von TNF80 und IL2 kloniert um die Bindungsspezifität des Fusionsproteins auf TNFR2, ebenso wie den IL2-Rezeptor welcher auf regulatorischen T-Zellen hoch exprimiert wird, herzustellen. Die spezifische Stimulation von Tregs würde der Therapie von Autoimmunerkrankungen dienen. In der Abteilung für Molekulare Innere Medizin in Würzburg wurde eine kovalent verknüpfte, nonamere Form von TNF, nämlich eine single-chain-TNF-TNC-Form hergestellt, sodass auch die Aktivierung von TNFR2 durch lösliches TNF möglich ist, was zur klinischen Anwendung (durch Injektionen) notwendig ist. Nach Klonierung und Produktion der Konstrukte in HEK293-Zellen erfolgte deren Aufreinigung und Quantifizierung. Letztendlich wurde mittels Bindungsstudien die Funktionalität der aufgereinigten Fusionsproteine überprüft. Zukünftige Studien müssen nun aufklären, ob die IL8-Produktion durch TNF80(h)-Flag-IL2(h) bzw. TNF80(mu)-Flag-IL2(mu) stimuliert wird, nachdem der IL2-Teil der Konstrukte den IL2-Rezeptor gebunden hat. N2 - The tumornekrosisfactor (TNF) is a cytocine which has a multifunctional role in the immune system. TNF can bind two receptors to activate different signaling pathways: TNFR1 (TNF receptor type 1) or TNFR2 (TNF receptor type 2). TNF occurs as a membrane-integrated form or as a soluble homotrimeric cytokine. TNFR1 can be activated by the membrane-bound and the soluble trimeric form, whereas TNFR2 can be acivated by the membran-bound form of TNF. For specific regulation and a better control of the TNF-binding TNF-mutants were developed. While TNF60 specifically binds TNFR1, TNF80 specifically binds TNFR2 only. Binding TNF80 to TNFR2 results in an expansion of regulatory T cells. Within the frame of this dissertation a TNFR2-specific agonist with the functional domain IL2 was developed and characterised, on the one hand, to achieve an expansion of regulatory T-cells. The specific stimulation of regulatory T-cells could be used as therapeutic medium for autoimmune disease. On the other hand, fusion proteins with TNF80 and the antibody fragment of IgG-Fc were created to achieve a depletion of regulatory T cells by provoking an antibody dependent cellular cytotoxicity (ADCC). This process could be used in therapy against cancer. KW - TNF KW - TNFR2-spezifische Agonisten KW - Klonierung KW - Entwicklung KW - Autoimmuntherapie KW - Antitumorforschung Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-155486 ER - TY - THES A1 - Lutz, Mathias T1 - T-Zell-Immunität gegen die tumorassoziierten Antigene HER2/neu, MUC1, PRAME und WT1 bei gesunden Blutspendern und Schwangeren als ein immuntherapeutisches Modell für die allogene Blutstammzelltransplantation T1 - T-cell immunity against the tumor-associated antigens HER2/neu, MUC1, PRAME and WT1 in healthy blood donors and pregnant women as an immunotherapeutic model for allogeneic blood stem cell transplantation N2 - Der Erfolg der allogenen hämatopoetischen Stammzelltransplantation (HSZT) als Immuntherapie basiert neben den Minorantigendifferenzen zwischen Spender und Empfänger entscheidend auf einer spendervermittelten Immunität gegen tumorassoziierte Antigene (TAA), über deren Herkunft und Frequenz bei gesunden Blutspendern die derzeitige Studienlage kaum Informationen bietet. Da für viele klinisch relevante TAA eine Expression im fetalen und plazentaren Gewebe bekannt ist, wurde in dieser Arbeit die Schwangerschaft als möglicher Auslöser dieser T-lymphozytären Gedächtnisimmunantworten im Sinne eines Tumor- und Transplantationsmodells untersucht. Hierfür wurden insgesamt 114 gesunde Blutspender in drei Subgruppen aus 38 Frauen mit negativer Schwangerschaftsanamnese, 38 Frauen mit positiver Geburtenanamnese und 38 Männern in einer Querschnittsstudie betrachtet, daneben wurden 44 Frauen longitudinal während und nach ihrer ersten Schwangerschaft untersucht. Dabei wurden die CD8-positiven T-Lymphozyten der Probanden isoliert, mit Peptiden der vier klinisch relevanten TAA HER2/neu (human epidermal growth factor receptor 2), MUC1 (Mucin 1), PRAME (preferentially expressed antigen of melanoma) und WT1 (Wilms tumor protein 1) stimuliert und die Produktion von IFN-γ-mRNA mittels RT-qPCR gemessen. Daneben wurden zum Vergleich durchflusszytometrische und ELISPOT-basierte Verfahren durchgeführt. Bei den gesunden Blutspendern konnten CD8-positive Gedächtnisimmunantworten von niedriger und/oder hoher funktioneller Avidität gegen alle vier untersuchten TAA gemessen werden: Die Frequenz der dabei als „positiv“ definierten Immunantworten betrug bei HER2/neu 5 %, bei MUC1 14 %, bei PRAME 7 % und bei WT1 15 %. Männer wiesen insgesamt höhere absolute Level der Immunantworten gegen die untersuchten TAA auf, was auf eine testikuläre Expression dieser Antigene zurückzuführen sein könnte. In der Longitudinalanalyse bei den erstschwangeren Frauen ließen sich die stärksten Immunantworten zu Beginn der Schwangerschaft nachweisen, so dass es hier zu einem „Boost“ präexistenter TAA-spezifischer Autoimmunität zu kommen scheint. Durch das immunsuppressive hormonelle Milieu im Verlauf der Schwangerschaft und den Verlust der Zielantigene der feto-plazentaren Einheit durch die Geburt und Nachgeburt scheint diese Immunität aber nicht zu persistieren. Dadurch erklärt sich auch die Beobachtung, dass Frauen mit einer positiven Geburtenanamnese keine stärkeren Immunantworten gegen die untersuchten TAA aufwiesen als Frauen mit einer negativen Schwangerschaftsanamnese. Die Schwangerschaft hinterlässt diesbezüglich also ohne die Anwesenheit der vermittelnden Antigene keinen regelhaft bleibenden Effekt. Diese Resultate decken sich mit Beobachtungen aus der Tumorimmuntherapie, bei denen Vakzinierungen gegen TAA zwar eine kurzfristige Immunität generieren konnten, die aber nicht persistierte. Im Rahmen der HSZT kann eine solche TAA-spezifische Immunität vom Spender auf den Empfänger transferiert werden und vermag dann aufgrund des proinflammatorischen Immunmilieus sehr wohl zu expandieren und in einem begrenzten Ausmaß auch zu persistieren. Dementsprechend ergeben sich aus den in dieser Arbeit gewonnenen Resultaten relevante Implikationen für die allogene und in geringerem Ausmaß die autologe HSZT, daneben aber auch für innovative Tumortherapien wie die Immuncheckpoint-Blockade, da die Persistenz von tumorspezifischer Immunität letztendlich hochrelevant für eine langfristige Tumorkontrolle und damit für ein tumorfreies Überleben ist. Das vorliegende Modell trägt somit zum Verständnis der komplexen immunregulatorischen Vorgänge bei der Tumorkontrolle bei. Ob die hierbei aufgezeigten Immunantworten generell zu einer verbesserten TAA-spezifischen Immunrekonstitution und konsekutiv zu einem besseren klinischen Ergebnis beitragen, bleibt offen und wird in klinischen Studien geklärt werden müssen. N2 - Efficacy of allogeneic hematopoietic stem cell transplantation (HSCT) as immunotherapy is mediated by donor-derived immunity against both minor histocompatibility antigens and tumor-associated antigens (TAAs). However, so far little is known about the origin and frequency of immunity against the latter in healthy blood donors. Since expression in fetal and placental tissues is known for many TAAs with clinical relevance, we investigated the pregnancy as a potential trigger for these memory-type T cell immune responses. In a cross-sectional analysis, we examined a total of 114 healthy blood donors including 38 nulligravidous women, 38 primi- or multiparous women and 38 men. In a longitudinal analysis, we investigated a total of 44 women at four time points during and after their first pregnancy. Donors’ positively isolated CD8-positive T cells were pulsed with peptides of four TAAs with known clinical relevance and placental expression: HER2/neu (human epidermal growth factor receptor 2), MUC1 (Mucin 1), PRAME (preferentially expressed antigen of melanoma) und WT1 (Wilms tumor protein 1). Quantitative reverse-transcription polymerase chain reaction was applied to detect interferon-γ (IFN-γ) mRNA after this stimulation. Additionally, CD8-positive T cells were analyzed by flow cytometry and IFN-γ ELISPOT assays. In healthy blood donors, CD8-positive memory-type T cell immune responses of low and/or high avidity could be detected against all four analyzed TAAs. Frequency of positive immune responses varied from 5 % (HER2/neu) and 7 % (PRAME) to 14 % (MUC1) and 15 % (WT1). Higher absolute levels of immune responses were found in healthy men, likely due to testicular expression of the analyzed antigens. In healthy primigravidous women, strongest immune responses could be shown during pregnancy to disappear after delivery and completion of nursing. Therefore, it can be assumed that expression of TAAs in the fetoplacental unit is leading to a “boost” of pre-existing immunity against these antigens. However, these immune responses seem to be short-lived due to both permissive hormonal environment evolving during pregnancy and loss of the fetoplacental unit as driving target after delivery. Accordingly, no association between history of prior deliveries and stronger immune responses against the analyzed TAAs was observed in this work. The results of our study are in line with observations made in tumor immunotherapy where immunity against TAAs could be generated by vaccination strategies, but showed no long-lasting persistence. Nevertheless, the shown TAA-specific immunity could be transferred into recipients after allogeneic HSCT, where a proinflammatory environment provides capability for homeostatic expansion and persistence of these immune responses. Taken together, the findings of this work provide implications for both allogeneic and autologous HSCT. Furthermore, our results could also gain relevance for innovative immunotherapies such as immune checkpoint inhibitors since persistence of tumor-specific immunity is highly important for control of the malignant disease and consecutively for long-term survival. Whether TAA-specific immune responses as shown in our study could contribute to an improved immune reconstitution after allogeneic HSCT and consecutively to a better long-term outcome remains unclear and needs to be clarified in clinical trials. KW - Immuntherapie KW - Periphere Stammzellentransplantation KW - Transplantat-Wirt-Reaktion KW - Tumorantigen KW - Schwangerschaft KW - Allogene hämatopoetische Stammzelltransplantation KW - Graft-versus-Leukämie-Effekt KW - Tumorassoziierte Antigene KW - immunotherapy KW - allogeneic hematopoietic stem cell transplantation KW - graft-versus-leukemia effect KW - tumor-associated antigens KW - pregnancy Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-156587 ER - TY - THES A1 - Müller, Nadine T1 - Nachweis von zirkulierenden Tumorzellen (CTC) in venösem Vollblut von Patienten mit gastrointestinalen Tumoren : Untersuchungen in vitro N2 - Zirkulierende Tumorzellen (CTCs) sind maligne Zellen, die in sehr geringer Anzahl im peripheren Blut von Tumorpatienten zu finden sind. Sie können entweder vom Primär-tumor oder von Metastasen in sekundären Organen stammen und sind in der Lage, sich nach dem Verlassen der Blutbahn in verschiedenen Geweben anzusiedeln und neue Me-tastasen auszubilden. Der Nachweis dieser CTCs im Blut ist mittlerweile zu einem viel-versprechenden und viel beforschtem Gebiet der Onkologie geworden. Das Vorhanden-sein von CTCs im Blut von Tumorpatienten kann ein früher Indikator für eine Metasta-sierung sein, als zuverlässiger Vorhersageparameter für die Prognose dienen und die Effektivität einer Therapie aufzeigen. In der vorliegenden Arbeit wurde ein immunologisches Verfahren entwickelt, womit CTCs in venösem Vollblut schnell und einfach als EpCAM-und Zytokeratin-positive und CD45-negative Zellen nachgewiesen werden können. Dabei erfolgt nach einer Ly-se der Erythrozyten in der Probe eine Leukozytendepletion mittels MACS (magnetic-activated cell sorting) und die Identifikation der Tumorzellen mittels FACS-Analyse mit einer festgelegten Gatingstrategie. Mit diesem Verfahren wurden Blutproben von 42 Patienten mit metastasierten gastrointestinalen Tumoren und von 10 gesunden Normal-spendern auf die Anzahl von CTCs/3,75ml Blut untersucht. Dabei wiesen die Patienten annähernd signifikant (p=0,076) mehr CTCs auf als die Normalspender. In 43% der Fäl-le hatten die Patienten >2 CTCs/3,75ml Vollblut und galten damit als CTC-positiv. Pati-enten mit Kolorektalem Karzinom zeigten mit 71% den höchsten Anteil an CTC-Positivität. Bei den gesunden Normalspendern ließen sich in keinem Fall >2 CTCs nachweisen. Nach einer Evaluation dieser Methode in einer weiteren Studie mit einem größeren Pati-enten- und Vergleichskollektiv könnte eine höhere Beweisebene erreicht werden, die den Einsatz in der klinischen Routine ermöglicht. Daneben sind eine weitere Optimie-rung des FACS-Verfahrens und eine Erweiterung des Antigenspektrums bei der CTC-Detektion zur Erhöhung der Sensitivität und Spezifität erforderlich. Patienten würden von diesem einfachen und wenig invasivem CTC-Detektionsverfahren vor allem im Hinblick auf eine Individualisierung der Therapie und auf das Vermeiden einer Überthe-rapie deutlich profitieren. N2 - Circulating tumor cells are malignant cells which can be found in very low number in the peripheral blood of patients with malignant diseases. They originate either from the primary tumor or from metastases in secondary organs and are able to settle down in different tissues and form new metastases after leaving circulation. Detection of CTCs in the blood has come to a promising and much researched field in oncology. The presence of CTCs in the blood of patients with malignant diseases can be an early indicator for metastasizing, serve as a reliable prediction parameter for prognosis and show the efficacy of therapy. In this paper an immunological method was developed to detect CTCs in venous whole blood fast and easily as EpCAM and cytokeratin positive and CD45 negative cells. In this method after lysis of erythrocytes in the sample a depletion of the leucocytes by MACS (magnetic activates cell sorting) and the identification of the tumor cells by FACS analysis with a determined gating strategy is accomplished. With this method blood samples of 42 patients with metastatic gastrointestinal tumors and 10 healthy donors were tested for the number of CTCs/3,75 ml blood. Patients had more CTCs than the healthy donors with approaching significance (p=0,076). 43% of the patients had > 2 CTCs/3,75 ml whole blood and were considered to be CTC positive. Patients with colorectal cancer showed with 71% the highest ratio of CTC positivity. In the group of healthy donors > 2 CTCs could not be detected. After evaluating this method in further studies with a bigger collective of patients and healthy donors evidence with higher level could be reached so that the use in clinical routine is possible. Therefore a further improvement of the FACS and an extension of the antigen spectrum within the CTC detection is necessary for increase of sensitivity and specificity. Patients would clearly profit from this easy and low invasive CTC detection method especially in terms of individualization of therapy and avoiding an overtherapy. T2 - Detection of circulating tumor cells (CTC) in venous whole blood of patients with gastrointestinal tumors KW - zirkulierende Tumorzellen KW - gastrointestinale Tumore KW - circulating tumor cells KW - gastrointestinal tumors KW - circulating Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-152546 ER - TY - THES A1 - Klessen, Katharina Carola T1 - Der Einfluss der Immunmodulatoren R848 und Alum auf die angeborene Immunantwort humaner dendritischer Zellen gegen Aspergillus fumigatus T1 - The influence of the immune modulators R848 and Alum on the innate immune Response of human dendritic cells against Aspergillus fumigatus N2 - Invasive Aspergillosen zählen auch heute noch zu den potentiell lebensbedrohlichen Infektionen, die gemeinsam mit anderen invasiven Pilzinfektionen für die hohe Mortalität bei immunsupprimierten Patienten verantwortlich sind (Lin et al. 2001). Die Entwicklung und Erforschung spezifischer diagnostischer Methoden und antimykotischer Medikamente konnten die Behandlungschancen einer IA zwar verbessern, bringen aber weiterhin keine befriedigenden Erfolge. So ist es dringend erforderlich, alternative Therapieoptionen zu erforschen und zu entwickeln. Da sich seit einigen Jahren das Augenmerk vermehrt in Richtung Immuntherapie konzentriert und diese Therapieform auch bei der Behandlung invasiver Aspergillosen Anwendung findet, wurden in diesem Zusammenhang die Immunmodulatoren Resiquimod und Alum auf ihre Wirkung auf dendritische Zellen bei einer Aspergillus-Infektion analysiert. Dendritische Zellen besitzen in der Immunabwehr gegen Aspergillus eine Schlüsselrolle, indem sie als Bindeglied zwischen adaptivem und angeborenem Immunsystem fungieren und somit essentiell für eine effektive T-Zell gesteuerte Immunantwort sind. Der mögliche Einfluss der beiden Modulatoren auf die Sekretion inflammatorischer Zytokine dendritischer Zellen wurde auf Protein-Ebene untersucht und die Modifikation der Expression bestimmter Oberflächenmarker als Reaktion auf Resiquimod analysiert. Es zeigte sich, dass das Adjuvans Alum dendritische Zellen in ihrer Immunantwort gegen Aspergillus nicht beeinflusst und zu keiner gesteigerten Sekretion inflammatorischer Zytokine führt. Aus diesem Grund wurde auf die Bestimmung des Expressionsmuster der Oberflächenmoleküle auf dendritischen Zellen in Abhängigkeit von Alum verzichtet. Hingegen konnte Resiquimod einen positiven Trend in der verstärkten Zytokinsekretion aufweisen. So ließ sich in Anwesenheit von Resiquimod eine verstärkte pro-inflammatorische Immunantwort gegen Aspergillus fumigatus erkennen. Dieser additive Effekt von R848 zeigte sich auch bei der Expression kostimulatorischer Moleküle dendritischer Zellen. Es zeigte sich eine gesteigerte Reifung pilzstimulierter dendritischer Zellen in Anwesenheit von Resiquimod durch Zunahme der Level von CD40, CD80 und CD86. In der Expression des Markers CD83 konnte keine einheitliche Aussage getroffen werden, da es spenderabhängig sowohl zu einer Zu-, als auch Abnahme der Fluoreszenzintensität von CD83 als Reaktion auf eine Ko-Stimulation mit Aspergillus und R848 kam. Es war festzustellen, dass die Zellen auf die eingesetzten Stimulantien stark spenderabhängig reagieren. Auf Grundlage dieser Ergebnisse könnte sich ein möglicher Nutzen des Immunmodulators Resiquimod für die Therapie invasiver Aspergillosen ergeben. Gerade immunsupprimierte Patienten mit einer invasiven Aspergillose könnten von einer DC-basierten Immuntherapie in Verbindung mit Resiquimod profitieren. Dies gilt es jedoch nur, wenn es durch weitere Analysen und Versuchsreihen bestätigt werden kann. N2 - Invasive aspergillosis is still one of the potentially life-threatening infections which together with other invasive infections are responsible for the high mortality in immunosuppressed patients. The development and investigation of specific diagnostic methods and antifungal drugs have improved the treatment options of an IA, but they still do not provide satisfactory results Successes. It is therefore urgently necessary to explore and develop alternative therapy options. Since for several years the focus has increasingly been concentrated on immunotherapy, and this form of therapy is also used in the treatment of invasive aspergillosis, the immunomodulators Resiquimod and Alum were analyzed for their effect on dendritic cells in Aspergillus infection. Dendritic cells have a key role to play in immune defense against Aspergillus by acting as a link between adaptive and innate immune systems and thus essential for an effective T-cell-directed immune response. The possible influence of the two modulators on the secretion of inflammatory cytokines of dendritic cells was investigated at the protein level and the modification of the expression of certain surface markers in response to resiquimod was analyzed. It was found that the adjuvant alum does not influence dendritic cells in their immune response against Aspergillus and leads to no increased secretion of inflammatory cytokines. For this reason, the determination of the expression pattern of the surface molecules on dendritic cells as a function of Alum was dispensed with. On the other hand, Resiquimod showed a positive trend in the increased cytokine secretion. An increased pro-inflammatory immune response against Aspergillus fumigatus was detected in the presence of resiquimod. This additive effect of R848 was also shown in the expression of costimulatory molecules of dendritic cells. There was an increased maturation of fungi-stimulated dendritic cells in the presence of resiquimod by increasing the levels of CD40, CD80 and CD86. In the expression of the marker CD83, no uniform statement could be made because it was donor-dependent both to an increase and decrease in the fluorescence intensity of CD83 in response to co-stimulation with Aspergillus and R848. It was found that the cells respond strongly to the stimulants used. On the basis of these results, a possible benefit of the immunomodulator resiquimod for the therapy of invasive aspergilloses could result. Immunosuppressed patients with invasive aspergillosis may benefit from DC-based immunotherapy in combination with resiquimod. However, this is only possible if it can be confirmed by further analyzes and trials. KW - Aspergillose KW - Resiquimod KW - Aluminiumhydroxide KW - Aspergillose KW - Resiquimod KW - Aluminiumhydroxid KW - Immunsystem Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-153907 ER - TY - JOUR A1 - Hefner, Jochen A1 - Berberich, Sara A1 - Lanvers, Elena A1 - Sanning, Maria A1 - Steimer, Ann-Kathrin A1 - Kunzmann, Volker T1 - New insights into frequency and contents of fear of cancer progression/recurrence (FOP/FCR) in outpatients with colorectal carcinoma (CRC) receiving oral capecitabine: a pilot study at a comprehensive cancer center JF - Patient Preference and Adherence N2 - Background: Fear of cancer progression/recurrence (FOP/FCR) is considered one of the most prevalent sources of distress in cancer survivors and associated with lower quality of life and functional impairment. Detailed measures of FOP/FCR are needed because little is known about the knowledge of FOP/FCR, its associations with the patient–doctor relationship, and the rate of adequate therapy. Colorectal cancer (CRC) is one of the most prevalent cancer entities, and oral capecitabine is widely prescribed as treatment. Therefore, we initiated a pilot study to expand the literature on FOP/FCR in CRC outpatients receiving capecitabine and to generate hypotheses for future investigations. Methods: This study included 58 patients treated at a comprehensive cancer center. FOP/FCR was assessed with the Fear of Progression Questionnaire (FOP-Q-SF). Satisfaction with the relationships with doctors was assessed with the Patient–Doctor Relationship Questionnaire-9 (PRDQ-9). Levels of side effects were rated by the patients on a visual analog scale. Clinical data were extracted from the charts. Results: A total of 19 out of 58 patients (36%) suffered from FOP/FCR according to our assessment. Levels of FOP/FCR seemed to be mostly moderate to high. Only four out of the 19 distressed patients (21%) were treated accordingly. Typical side effects of oncological treatment were associated with higher FOP/FCR. Satisfaction with doctor–patient relationships was not associated with FOP/FCR. Regarding single items of FOP/FCR, three out of the five most prevalent fears were associated with close relatives. Discussion: FOP/FCR occurred frequently in more than one in three patients, but was mostly untreated in this sample of consecutive outpatients with CRC receiving oral capecitabine. In detail, most fears were related to family and friends. In addition to an unmet need of patients, our data indicate sources of distress not considered thus far. If replicated in larger studies, results may help to inform intervention development and improve patient care. KW - fear of progression KW - comprehensive management KW - oral anticancer drugs KW - colorectal cancer KW - screening for distress Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158476 VL - 11 ER - TY - THES A1 - Garcia Guerrero, Estefania T1 - Strategies to Obtain Tumor-Reactive Cells for Cancer Immunotherapy by Cell Sorting and Genetic Modifications of T Lymphocytes T1 - Zellsortierung und genetische Modifikation von T-Lymphozyten zur Gewinnung tumorreaktiver Zellen für die Krebsimmuntherapie N2 - Recent advances in the field of cancer immunotherapy have enabled this therapeutic approach to enter the mainstream of modern cancer treatment. In particular, adoptive T cell therapy (ACT) is a potentially powerful immunotherapy approach that relies on the administration of tumor-specific T cells into the patient. There are several strategies to obtain tumor-reactive cytotoxic T lymphocytes (CTLs), which have already been shown to induce remarkable responses in the clinical setting. However, there are concerns and limitations regarding the conventional approaches to obtain tumor-reactive T cells, such as accuracy of the procedure and reproducibility. Therefore, we aimed to develop two approaches to improve the precision and efficacy of tumor-reactive T cells therapy. These two techniques could constitute effective, safe and broadly applicable alternatives to the conventional methods for obtaining tumor-specific CTLs. The first approach of this study is the so called “Doublet Technology”. Here, we demonstrate that peptide-human leukocyte antigen-T cell receptor (pHLA-TCR) interactions that involve immune reactive peptides are stable and strong. Therefore, the CTLs that are bound by their TCR to tumor cells can be selected and isolated through FACS-based cell sorting taking advantage of this stable interaction between the CTLs and the target cells. The CTLs from acute myeloid leukemia (AML) patients obtained with this technique show cytolytic activity against blast cells suggesting a potential clinical use of these CTLs. “Doublet Technology” offers a personalized therapy in which there is no need for a priori knowledge of the exact tumor antigen. The second approach of this study is the Chimeric Antigen Receptor (CAR) Technology. We design several CARs targeting the B-Cell Maturation Antigen (BCMA). BCMA CAR T cells show antigen-specific cytolytic activity, production of cytokines including IFN-γ and IL-2, as well as productive proliferation. Although we confirm the presence of soluble BCMA in serum of multiple myeloma (MM) patients, we demonstrate that the presence of soluble protein does not abrogate the efficacy of BCMA CAR T cells suggesting that BCMA CAR T cells can be used in the clinical setting to treat MM patients. The high antigen specificity of CAR T cells allows efficient tumor cell eradication and makes CAR Technology attractive for broadly applicable therapies. N2 - Durch jüngste Fortschritte auf dem Gebiet der Krebsimmuntherapie konnte dieser therapeutische Ansatz in der Mitte moderner Krebsbehandlungen ankommen. Insbesondere die adoptive T-Zelltherapie (ACT), die auf der Verabreichung tumorspezifischer T-Zellen an den Patienten beruht, stellt einen potentiell schlagkräftigen immuntherapeutischen Ansatz dar. Es existieren bereits verschiedene Strategien um tumorreaktive zytotoxische T-Lymphozyten (CTL) herzustellen, von denen bereits gezeigt wurde, dass sie klinisch bemerkenswerte Antworten hervorrufen. Dennoch gibt es Bedenken und Grenzen bezüglich dieser konventionellen Ansätze zur Herstellung tumorreaktiver T-Zellen, wie zum Beispiel die Genauigkeit und Reproduzierbarkeit des Verfahrens. Daher arbeiteten wir an der Entwicklung zweier Ansätze um die Präzision und Effizienz der tumorreaktiven T-Zelltherapie zu verbessern. Diese beiden Techniken könnten effektive, sichere und breit anwendbare Alternativen zu den konventionellen Methoden der tumorspezifischen CTL-Gewinnung darstellen. Der erste Ansatz dieser Studie wird als „Doublet Technology“ bezeichnet. Hierbei zeigen wir, dass die Interaktionen zwischen Peptid/MHC-Komplex und T-Zellrezeptor (pHLA-TCR), die immunreaktive Peptide involvieren, stabil und solide sind. Außerdem zeigen wir, dass CTLs, die über ihren TCR an Tumorzellen gebunden sind, selektioniert und durch FACS-basierte Zellsortierung isoliert werden können. Hierbei wird die Stabilität der Interaktion von CTLs und Zielzellen genutzt. Die CTLs von Patienten mit Akuter Myeloischer Leukämie (AML), die auf diese Weise gewonnen werden, zeigen zytolytische Aktivität gegenüber Blasten, was auf einen potentiellen klinischen Nutzen dieser CTLs hinweisen könnte. Die „Doublet Technology“ bietet eine personalisierte Therapie, die kein vorheriges Wissen über ein exaktes Tumorantigen erfordert. Der zweite Ansatz dieser Studie ist die Chimere Antigenrezeptor (CAR) Technologie. Wir entwickeln verschiedene CARs gegen das B-Zellmaturationsantigen (BCMA). BCMA-CAR T-Zellen zeigen antigenspezifische zytolytische Aktivität, Produktion der Zytokine IFN-γ und IL-2 sowie produktive Proliferation. Obwohl wir bestätigen, dass lösliches BCMA im Serum von Multiplen Myelompatienten zu finden ist, zeigen wir auch, dass dieses lösliche Protein nicht die Effizienz von BCMA-CAR T-Zellen beeinträchtigt und somit BCMA-CAR T-Zellen zur Behandlung von Multiplen Myelompatienten klinisch genutzt werden können. Die hohe Antigenspezifität der CAR-T-Zellen erlaubt eine effiziente Vernichtung von Tumorzellen und macht die CAR-Technologie attraktiv für breit einsetzbare Therapien. KW - Immunotherapy KW - Cancer KW - Strategies to Obtain Tumor-Reactive Cells Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-150547 ER - TY - JOUR A1 - Neuchel, Christine A1 - Fürst, Daniel A1 - Niederwieser, Dietger A1 - Bunjes, Donald A1 - Tsamadou, Chrysanthi A1 - Wulf, Gerald A1 - Pfreundschuh, Michael A1 - Wagner, Eva A1 - Stuhler, Gernot A1 - Einsele, Hermann A1 - Schrezenmeier, Hubert A1 - Mytilineos, Joannis T1 - Impact of donor activating KIR genes on HSCT outcome in C1-ligand negative myeloid disease patients transplanted with unrelated donors - a retrospective study JF - PLOS One N2 - Natural Killer cells (NK) are lymphocytes with the potential to recognize and lyse cells which escaped T-cell mediated lysis due to their aberrant HLA expression profiles. Killer cell immunoglobulin-like receptors (KIR) influence NK-cell activity by mediation of activating or inhibitory signals upon interaction with HLA-C (C1, C2) ligands. Therefore, absence of ligands for donor inhibitory KIRs following hematopoietic stem cell transplantation (HSCT) may have an influence on its outcome. Previous studies showed that C1 negative patients have a decreased HSCT outcome. Our study, based on a cohort of 200 C1-negative patients, confirmed these findings for the endpoints: overall survival (OS: HR = 1.41, CI = 1.14–1.74, p = 0.0012), disease free survival (DFS: HR = 1.27, CI = 1.05–1.53, p = 0.015), treatment related mortality (TRM: HR = 1.41, CI = 1.01–1.96, p = 0.04), and relapse incidence (RI: HR = 1.33, CI = 1.01–1.75, p = 0.04) all being inferior when compared to C1-positive patients (n = 1246). Subsequent analysis showed that these findings applied for patients with myeloid malignancies but not for patients with lymphoproliferative diseases (OS: myeloid: HR = 1.51, CI = 1.15–1.99, p = 0.003; lymphoblastic: HR = 1.26, CI = 0.91–1.75, p = 0.16; DFS: myeloid: HR = 1.31, CI = 1.01–1.70, p = 0.04; lymphoblastic: HR = 1.21, CI = 0.90–1.61, p = 0.21; RI: myeloid: HR = 1.31, CI = 1.01–1.70, p = 0.04; lymphoblastic: HR = 1.21, CI = 0.90–1.61, p = 0.21). Interestingly, within the C1-negative patient group, transplantation with KIR2DS2 resulted in better OS (9/10 matched: HR = 0.24, CI = 0.08–0.67, p = 0.007) as well as DFS (9/10 matched: HR = 0,26, CI = 0.11–0.60, p = 0.002), and transplantation with KIR2DS1 positive donors was associated with a decreased RI (HR = 0.30, CI = 0.13–0.69, p = 0.005). TRM was increased when the donor was positive for KIR2DS1 (HR = 2.61, CI = 1.26–5.41, p = 0.001). Our findings suggest that inclusion of KIR2DS1/2/5 and KIR3DS1-genotyping in the unrelated donor search algorithm of C1-ligand negative patients with myeloid malignancies may prove to be of clinical relevance. KW - Hematopoietic stem cell transplantation KW - Cancer risk factors KW - Multivariate analysis KW - Stem cell transplantation KW - T-cells KW - Bone marrow transplantation KW - NK-cells KW - Immune receptor signaling KW - Killer cell immunoglobulin-like receptors KW - Acute myeloid leukemia KW - Acute lymphoblastic leukemia KW - Chronic lymphoblastic leukemia KW - Chronic myeloid leukemia Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-180995 VL - 12 IS - 1 ER - TY - JOUR A1 - Werner, Rudolf A. A1 - Weich, Alexander A1 - Higuchi, Takahiro A1 - Schmid, Jan S. A1 - Schirbel, Andreas A1 - Lassmann, Michael A1 - Wild, Vanessa A1 - Rudelius, Martina A1 - Kudlich, Theodor A1 - Herrmann, Ken A1 - Scheurlen, Michael A1 - Buck, Andreas K. A1 - Kropf, Saskia A1 - Wester, Hans-Jürgen A1 - Lapa, Constantin T1 - Imaging of Chemokine Receptor 4 Expression in Neuroendocrine Tumors - a Triple Tracer Comparative Approach JF - Theranostics N2 - C-X-C motif chemokine receptor 4 (CXCR4) and somatostatin receptors (SSTR) are overexpressed in gastro-entero-pancreatic neuroendocrine tumors (GEP-NET). In this study, we aimed to elucidate the feasibility of non-invasive CXCR4 positron emission tomography/computed tomography (PET/CT) imaging in GEP-NET patients using [\(^{68}\)Ga]Pentixafor in comparison to \(^{68}\)Ga-DOTA-D-Phe-Tyr3-octreotide ([\(^{68}\)Ga]DOTATOC) and \(^{18}\)F-fluorodeoxyglucose ([\(^{18}\)F]FDG). Twelve patients with histologically proven GEP-NET (3xG1, 4xG2, 5xG3) underwent [\(^{68}\)Ga]DOTATOC, [\(^{18}\)F]FDG, and [\(^{68}\)Ga]Pentixafor PET/CT for staging and planning of the therapeutic management. Scans were analyzed on a patient as well as on a lesion basis and compared to immunohistochemical staining patterns of CXCR4 and somatostatin receptors SSTR2a and SSTR5. [\(^{68}\)Ga]Pentixafor visualized tumor lesions in 6/12 subjects, whereas [\(^{18}\)F]FDG revealed sites of disease in 10/12 and [\(^{68}\)Ga]DOTATOC in 11/12 patients, respectively. Regarding sensitivity, SSTR-directed PET was the superior imaging modality in all G1 and G2 NET. CXCR4-directed PET was negative in all G1 NET. In contrast, 50% of G2 and 80% of G3 patients exhibited [\(^{68}\)Ga]Pentixafor-positive tumor lesions. Whereas CXCR4 seems to play only a limited role in detecting well-differentiated NET, increasing receptor expression could be non-invasively observed with increasing tumor grade. Thus, [\(^{68}\)Ga]Pentixafor PET/CT might serve as non-invasive read-out for evaluating the possibility of CXCR4-directed endoradiotherapy in advanced dedifferentiated SSTR-negative tumors. KW - SSTR KW - peptide receptor radionuclide therapy KW - neuroendocrine tumor KW - [\(^{68}\)Ga]Pentixafor KW - CXCR4 KW - chemokine receptor KW - PET/CT KW - DOTATOC KW - PRRT KW - Positronen-Emissions-Tomografie Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158008 VL - 7 IS - 6 ER - TY - JOUR A1 - Wolter, Patrick A1 - Hanselmann, Steffen A1 - Pattschull, Grit A1 - Schruf, Eva A1 - Gaubatz, Stefan T1 - Central spindle proteins and mitotic kinesins are direct transcriptional targets of MuvB, B-MYB and FOXM1 in breast cancer cell lines and are potential targets for therapy JF - Oncotarget N2 - The MuvB multiprotein complex, together with B-MYB and FOXM1 (MMB-FOXM1), plays an essential role in cell cycle progression by regulating the transcription of genes required for mitosis and cytokinesis. In many tumors, B-MYB and FOXM1 are overexpressed as part of the proliferation signature. However, the transcriptional targets that are important for oncogenesis have not been identified. Given that mitotic kinesins are highly expressed in cancer cells and that selected kinesins have been reported as target genes of MMB-FOXM1, we sought to determine which mitotic kinesins are directly regulated by MMB-FOXM1. We demonstrate that six mitotic kinesins and two microtubule-associated non-motor proteins (MAPs) CEP55 and PRC1 are direct transcriptional targets of MuvB, B-MYB and FOXM1 in breast cancer cells. Suppression of KIF23 and PRC1 strongly suppressed proliferation of MDA-MB-231 cells. The set of MMB-FOXM1 regulated kinesins genes and 4 additional kinesins which we referred to as the mitotic kinesin signature (MKS) is linked to poor outcome in breast cancer patients. Thus, mitotic kinesins could be used as prognostic biomarker and could be potential therapeutic targets for the treatment of breast cancer. KW - breast cancer KW - kinesin KW - cell cycle KW - cytokinesis Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-171851 VL - 8 IS - 7 ER - TY - JOUR A1 - Sander, Bodo A1 - Xu, Wenshan A1 - Eilers, Martin A1 - Popov, Nikita A1 - Lorenz, Sonja T1 - A conformational switch regulates the ubiquitin ligase HUWE1 JF - eLife N2 - The human ubiquitin ligase HUWE1 has key roles in tumorigenesis, yet it is unkown how its activity is regulated. We present the crystal structure of a C-terminal part of HUWE1, including the catalytic domain, and reveal an asymmetric auto-inhibited dimer. We show that HUWE1 dimerizes in solution and self-associates in cells, and that both occurs through the crystallographic dimer interface. We demonstrate that HUWE1 is inhibited in cells and that it can be activated by disruption of the dimer interface. We identify a conserved segment in HUWE1 that counteracts dimer formation by associating with the dimerization region intramolecularly. Our studies reveal, intriguingly, that the tumor suppressor p14ARF binds to this segment and may thus shift the conformational equilibrium of HUWE1 toward the inactive state. We propose a model, in which the activity of HUWE1 underlies conformational control in response to physiological cues—a mechanism that may be exploited for cancer therapy. KW - Medicine KW - Structural Biology KW - Molecular Biophysics KW - HUWE1 KW - HECT Ligase KW - Ubiquitin KW - P14ARF KW - X-Ray Chrystallography KW - Enzyme Regulation Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-171862 VL - 6 ER - TY - JOUR A1 - Lapa, Constantin A1 - Herrmann, Ken A1 - Schirbel, Andreas A1 - Hänscheid, Heribert A1 - Lückerath, Katharina A1 - Schottelius, Margret A1 - Kircher, Malte A1 - Werner, Rudolf A. A1 - Schreder, Martin A1 - Samnick, Samuel A1 - Kropf, Saskia A1 - Knop, Stefan A1 - Buck, Andreas K. A1 - Einsele, Hermann A1 - Wester, Hans-Juergen A1 - Kortüm, K. Martin T1 - CXCR4-directed endoradiotherapy induces high response rates in extramedullary relapsed multiple myeloma JF - Theranostics N2 - C-X-C-motif chemokine receptor 4 (CXCR4) is a key factor for tumor growth and metastasis in several types of human cancer. We have recently reported promising first-in-man experience with CXCR4-directed endoradiotherapy (ERT) in multiple myeloma (MM). Eight heavily pretreated MM patients underwent a total of 10 ERT cycles (7 patients with 1 cycle and a single patient with 3 cycles). ERT was administered in combination with chemotherapy and autologous stem cell support. End points were occurrence and timing of adverse events, progression-free and overall survival. ERT was overall well tolerated without any unexpected acute adverse events or changes in vital signs. With absorbed tumor doses >30-70 Gy in intra- or extramedullary lesions, significant anti-myeloma activity was observed with 1 patient achieving complete remission and 5/8 partial remission. Directly after ERT major infectious complications were seen in one patient who died from sepsis 22 days after ERT, another patient with high tumor burden experienced lethal tumor lysis syndrome. Median progression-free survival was 54 days (range, 13-175), median overall survival was 223 days (range, 13-313). During follow-up (6 patients available), one patient died from infectious complications, 2/8 from disease progression, the remaining 3/8 patients are still alive. CXCR4-directed ERT was well-tolerated and exerted anti-myeloma activity even at very advanced stage MM with presence of extramedullary disease. Further assessment of this novel treatment option is highly warranted. KW - medicine KW - multiple myeloma KW - PET KW - CXCR4 KW - theranostics Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-172095 VL - 7 IS - 6 ER - TY - JOUR A1 - Lapa, Constantin A1 - Schreder, Martin A1 - Schirbel, Andreas A1 - Samnick, Samuel A1 - Kortüm, Klaus Martin A1 - Herrmann, Ken A1 - Kropf, Saskia A1 - Einsele, Herrmann A1 - Buck, Andreas K. A1 - Wester, Hans-Jürgen A1 - Knop, Stefan A1 - Lückerath, Katharina T1 - [\(^{68}\)Ga]Pentixafor-PET/CT for imaging of chemokine receptor CXCR4 expression in multiple myeloma - comparison to [\(^{18}\)F]FDG and laboratory values JF - Theranostics N2 - Chemokine (C-X-C motif) receptor 4 (CXCR4) is a key factor for tumor growth and metastasis in several types of human cancer including multiple myeloma (MM). Proof-of-concept of CXCR4-directed radionuclide therapy in MM has recently been reported. This study assessed the diagnostic performance of the CXCR4-directed radiotracer [\(^{68}\)Ga]Pentixafor in MM and a potential role for stratifying patients to CXCR4-directed therapies. Thirty-five patients with MM underwent [\(^{68}\)Ga]Pentixafor-PET/CT for evaluation of eligibility for endoradiotherapy. In 19/35 cases, [\(^{18}\)F]FDG-PET/CT for correlation was available. Scans were compared on a patient and on a lesion basis. Tracer uptake was correlated with standard clinical parameters of disease activity. [\(^{68}\)Ga]Pentixafor-PET detected CXCR4-positive disease in 23/35 subjects (66%). CXCR4-positivity at PET was independent from myeloma subtypes, cytogenetics or any serological parameters and turned out as a negative prognostic factor. In the 19 patients in whom a comparison to [\(^{18}\)F]FDG was available, [\(^{68}\)Ga]Pentixafor-PET detected more lesions in 4/19 (21%) subjects, [\(^{18}\)F]FDG proved superior in 7/19 (37%). In the remaining 8/19 (42%) patients, both tracers detected an equal number of lesions. [\(^{18}\)F]FDG-PET positivity correlated with [\(^{68}\)Ga]Pentixafor-PET positivity (p=0.018). [\(^{68}\)Ga]Pentixafor-PET provides further evidence that CXCR4 expression frequently occurs in advanced multiple myeloma, representing a negative prognostic factor and a potential target for myeloma specific treatment. However, selecting patients for CXCR4 directed therapies and prognostic stratification seem to be more relevant clinical applications for this novel imaging modality, rather than diagnostic imaging of myeloma. KW - medicine KW - multiple myeloma KW - FDG KW - molecular imaging KW - CXCR4 KW - PET KW - radionuclide therapy KW - theranostics Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-172106 VL - 7 IS - 1 ER - TY - JOUR A1 - Tsamadou, Chrysanthi A1 - Fürst, Daniel A1 - Vucinic, Vladan A1 - Bunjes, Donald A1 - Neuchel, Christine A1 - Mytilineos, Daphne A1 - Gramatzki, Martin A1 - Arnold, Renate A1 - Wagner, Eva Maria A1 - Einsele, Hermann A1 - Müller, Carlheinz A1 - Schrezenmeier, Hubert A1 - Mytilineos, Joannis T1 - Human leukocyte antigen-E mismatch is associated with better hematopoietic stem cell transplantation outcome in acute leukemia patients JF - Haematologica N2 - The immunomodulatory role of human leukocyte antigen (HLA)-E in hematopoietic stem cell transplantation (HSCT) has not been extensively investigated. To this end, we genotyped 509 10/10 HLA unrelated transplant pairs for HLA-E, in order to study the effect of HLA-E as a natural killer (NK)-alloreactivity mediator on HSCT outcome in an acute leukemia (AL) setting. Overall survival (OS), disease free survival (DFS), relapse incidence (RI) and non-relapse mortality (NRM) were set as endpoints. Analysis of our data revealed a significant correlation between HLA-E mismatch and improved HSCT outcome, as shown by both univariate (53% vs. 38%, P=0.002, 5-year OS) and multivariate (hazard ratio (HR)=0.63, confidence interval (CI) 95%=0.48–0.83, P=0.001) analyses. Further subgroup analysis demonstrated that the positive effect of HLA-E mismatch was significant and pronounced in advanced disease patients (n=120) (5-year OS: 50% vs. 18%, P=0.005; HR=0.40, CI 95%=0.22–0.72, P=0.002; results from univariate and multivariate analyses, respectively). The study herein is the first to report an association between HLA-E incompatibility and improved post–transplant prognosis in AL patients who have undergone matched unrelated HSCT. Combined NK and T cell HLA-E-mediated mechanisms may account for the better outcomes observed. Notwithstanding the necessity for in vitro and confirmational studies, our findings highlight the clinical relevance of HLA-E matching and strongly support prospective HLA-E screening upon donor selection for matched AL unrelated HSCTs. KW - medicine KW - acute leukemia (AL) KW - hematopoietic stem cell transplantation (HSCT) KW - human leukocyte antigen-E (HLA-E) KW - HLA-E matching KW - HSTC outcome Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-173325 VL - 102 IS - 11 ER - TY - JOUR A1 - Klein-Hessling, Stefan A1 - Muhammad, Khalid A1 - Klein, Matthias A1 - Pusch, Tobias A1 - Rudolf, Ronald A1 - Flöter, Jessica A1 - Qureischi, Musga A1 - Beilhack, Andreas A1 - Vaeth, Martin A1 - Kummerow, Carsten A1 - Backes, Christian A1 - Schoppmeyer, Rouven A1 - Hahn, Ulrike A1 - Hoth, Markus A1 - Bopp, Tobias A1 - Berberich-Siebelt, Friederike A1 - Patra, Amiya A1 - Avots, Andris A1 - Müller, Nora A1 - Schulze, Almut A1 - Serfling, Edgar T1 - NFATc1 controls the cytotoxicity of CD8\(^{+}\) T cells JF - Nature Communications N2 - Cytotoxic T lymphocytes are effector CD8\(^{+}\) T cells that eradicate infected and malignant cells. Here we show that the transcription factor NFATc1 controls the cytotoxicity of mouse cytotoxic T lymphocytes. Activation of Nfatc1\(^{-/-}\) cytotoxic T lymphocytes showed a defective cytoskeleton organization and recruitment of cytosolic organelles to immunological synapses. These cells have reduced cytotoxicity against tumor cells, and mice with NFATc1-deficient T cells are defective in controlling Listeria infection. Transcriptome analysis shows diminished RNA levels of numerous genes in Nfatc1\(^{-/-}\) CD8\(^{+}\) T cells, including Tbx21, Gzmb and genes encoding cytokines and chemokines, and genes controlling glycolysis. Nfatc1\(^{-/-}\), but not Nfatc2\(^{-/-}\) CD8\(^{+}\) T cells have an impaired metabolic switch to glycolysis, which can be restored by IL-2. Genome-wide ChIP-seq shows that NFATc1 binds many genes that control cytotoxic T lymphocyte activity. Together these data indicate that NFATc1 is an important regulator of cytotoxic T lymphocyte effector functions. KW - cytotoxic T cells KW - lymphocyte activation KW - signal transduction KW - gene regulation KW - immune cells KW - NFATc1 Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-170353 VL - 8 IS - 511 ER - TY - JOUR A1 - Hellmann, Anna-Maria A1 - Lother, Jasmin A1 - Wurster, Sebastian A1 - Lutz, Manfred B. A1 - Schmitt, Anna Lena A1 - Morton, Charles Oliver A1 - Eyrich, Matthias A1 - Czakai, Kristin A1 - Einsele, Hermann A1 - Loeffler, Juergen T1 - Human and Murine Innate Immune Cell Populations Display Common and Distinct Response Patterns during Their In Vitro Interaction with the Pathogenic Mold Aspergillus fumigatus JF - Frontiers in Immunology N2 - Aspergillus fumigatus is the main cause of invasive fungal infections occurring almost exclusively in immunocompromised patients. An improved understanding of the initial innate immune response is key to the development of better diagnostic tools and new treatment options. Mice are commonly used to study immune defense mechanisms during the infection of the mammalian host with A. fumigatus. However, little is known about functional differences between the human and murine immune response against this fungal pathogen. Thus, we performed a comparative functional analysis of human and murine dendritic cells (DCs), macrophages, and polymorphonuclear cells (PMNs) using standardized and reproducible working conditions, laboratory protocols, and readout assays. A. fumigatus did not provoke identical responses in murine and human immune cells but rather initiated relatively specific responses. While human DCs showed a significantly stronger upregulation of their maturation markers and major histocompatibility complex molecules and phagocytosed A. fumigatus more efficiently compared to their murine counterparts, murine PMNs and macrophages exhibited a significantly stronger release of reactive oxygen species after exposure to A. fumigatus. For all studied cell types, human and murine samples differed in their cytokine response to conidia or germ tubes of A. fumigatus. Furthermore, Dectin-1 showed inverse expression patterns on human and murine DCs after fungal stimulation. These specific differences should be carefully considered and highlight potential limitations in the transferability of murine host–pathogen interaction studies. KW - murine model KW - humans KW - Aspergillus fumigatus KW - innate immune response KW - fungal infection Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-169926 VL - 8 IS - 1716 ER - TY - JOUR A1 - Lapa, Constantin A1 - Garcia-Velloso, Maria J. A1 - Lückerath, Katharina A1 - Samnick, Samuel A1 - Schreder, Martin A1 - Otero, Paula Rodriguez A1 - Schmid, Jan-Stefan A1 - Herrmann, Ken A1 - Knop, Stefan A1 - Buck, Andreas K. A1 - Einsele, Hermann A1 - San-Miguel, Jesus A1 - Kortüm, Klaus Martin T1 - \(^{11}\)C-methionine-PET in multiple myeloma: a combined study from two different institutions JF - Theranostics N2 - \(^{11}\)C-methionine (MET) has recently emerged as an accurate marker of tumor burden and disease activity in patients with multiple myeloma (MM). This dual-center study aimed at further corroboration of the superiority of MET as positron emission tomography (PET) tracer for staging and re-staging MM, as compared to \(^{18}\)F-2`-deoxy-2`-fluoro-D-glucose (FDG). 78 patients with a history of solitary plasmacytoma (n=4), smoldering MM (SMM, n=5), and symptomatic MM (n=69) underwent both MET- and FDG-PET/computed tomography (CT) at the University Centers of Würzburg, Germany and Navarra, Spain. Scans were compared on a patient and on a lesion basis. Inter-reader agreement was also evaluated. In 2 patients, tumor biopsies for verification of discordant imaging results were available. MET-PET detected focal lesions (FL) in 59/78 subjects (75.6%), whereas FDG-PET/CT showed lesions in only 47 patients (60.3%; p<0.01), accordingly disease activity would have been missed in 12 patients. Directed biopsies of discordant results confirmed MET-PET/CT results in both cases. MET depicted more FL in 44 patients (56.4%; p<0.01), whereas in two patients (2/78), FDG proved superior. In the remainder (41.0%, 32/78), both tracers yielded comparable results. Inter-reader agreement for MET was higher than for FDG (κ = 0.82 vs κ = 0.72). This study demonstrates higher sensitivity of MET in comparison to standard FDG to detect intra- and extramedullary MM including histologic evidence of FDG-negative, viable disease exclusively detectable by MET-PET/CT. MET holds the potential to replace FDG as functional imaging standard for staging and re-staging of MM. KW - medicine KW - PET/CT KW - \(^{11}\)C-methionine KW - multiple myeloma KW - FDG Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-172038 VL - 7 IS - 11 ER - TY - JOUR A1 - Kelm, M. A1 - Seyfried, F. A1 - Reimer, S. A1 - Krajinovic, K. A1 - Miras, A. D. A1 - Jurowich, C. A1 - Germer, C. T. A1 - Brand, M. T1 - Proximal jejunal stoma as ultima ratio in case of traumatic distal duodenal perforation facilitating successful EndoVAC\(^{®}\) treatment: a case report JF - International Journal of Surgery Case Reports N2 - Introduction: During damage control surgery for blunt abdominal traumata simultaneous duodenal perforations can be missed making secondary sufficient surgical treatment challenging. Endoluminal vacuum (EndoVAC™) therapy has been shown to be a revolutionary option but has anatomical and technical limits. Presentation of the case: A 59-year old man with hemorrhagic shock due to rupture of the mesenteric root after blunt abdominal trauma received damage control treatment. Within a scheduled second-look, perforation of the posterior duodenal wall was identified. Due to local and systemic conditions, further surgical treatment was limited. Decision for endoscopic treatment was made but proved to be difficult due to the distal location. Finally, double-barreled jejunal stoma was created for transstomal EndoVAC™ treatment. Complete leakage healing was achieved and jejunostomy reversal followed subsequently. Discussion: During damage control surgery simultaneous bowel injuries can be missed leading to life-threatening complications with limited surgical options. EndoVAC™ treatment is an option for gastrointestinal perforations but has anatomical limitations that can be sufficiently shifted by a transstomal approach for intestinal leakage. Conclusion: In trauma related laparotomy complete mobilization of the duodenum is crucial. As ultima ratio, transstomal EndoVAC™ is a safe and feasible option and can be considered for similar cases. KW - transstomal endoluminal vacuum therapy KW - EndoVAC and small bowel KW - duodenal trauma KW - duodenal perforation Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-159292 VL - 41 ER - TY - JOUR A1 - Stegner, David A1 - van Eeuwijk, Judith M.M. A1 - Angay, Oğuzhan A1 - Gorelashvili, Maximilian G. A1 - Semeniak, Daniela A1 - Pinnecker, Jürgen A1 - Schmithausen, Patrick A1 - Meyer, Imke A1 - Friedrich, Mike A1 - Dütting, Sebastian A1 - Brede, Christian A1 - Beilhack, Andreas A1 - Schulze, Harald A1 - Nieswandt, Bernhard A1 - Heinze, Katrin G. T1 - Thrombopoiesis is spatially regulated by the bone marrow vasculature JF - Nature Communications N2 - In mammals, megakaryocytes (MKs) in the bone marrow (BM) produce blood platelets, required for hemostasis and thrombosis. MKs originate from hematopoietic stem cells and are thought to migrate from an endosteal niche towards the vascular sinusoids during their maturation. Through imaging of MKs in the intact BM, here we show that MKs can be found within the entire BM, without a bias towards bone-distant regions. By combining in vivo two-photon microscopy and in situ light-sheet fluorescence microscopy with computational simulations, we reveal surprisingly slow MK migration, limited intervascular space, and a vessel-biased MK pool. These data challenge the current thrombopoiesis model of MK migration and support a modified model, where MKs at sinusoids are replenished by sinusoidal precursors rather than cells from a distant periostic niche. As MKs do not need to migrate to reach the vessel, therapies to increase MK numbers might be sufficient to raise platelet counts. KW - bone marrow KW - megakaryocytes KW - thrombopoiesis Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-170591 VL - 8 IS - 127 ER - TY - JOUR A1 - Hergovits, Sabine A1 - Mais, Christine A1 - Haan, Claude A1 - Costa-Pereira, Ana P. A1 - Hermanns, Heike M. T1 - Oncostatin M induces RIG-I and MDA5 expression and enhances the double-stranded RNA response in fibroblasts JF - Journal of Cellular and Molecular Medicine N2 - Interleukin (IL)-6-type cytokines have no direct antiviral activity; nevertheless, they display immune-modulatory functions. Oncostatin M (OSM), a member of the IL-6 family, has recently been shown to induce a distinct number of classical interferon stimulated genes (ISG). Most of them are involved in antigen processing and presentation. However, induction of retinoic acid-inducible gene (RIG)-I-like receptors (RLR) has not been investigated. Here we report that OSM has the capability to induce the expression of the DExD/H-Box RNA helicases RIG-I and melanoma differentiation antigen 5 (MDA5) as well as of the transcription factors interferon regulatory factor (IRF)1, IRF7 and IRF9 in primary fibroblasts. Induction of the helicases depends on tyrosine as well as serine phosphorylation of STAT1. Moreover, we could show that the OSM-induced STAT1 phosphorylation is predominantly counter-regulated by a strong STAT3-dependent SOCS3 induction, as Stat3 as well as Socs3 knock-down results in an enhanced and prolonged helicase and IRF expression. Other factors involved in regulation of STAT1 or IRF1 activity, like protein tyrosine phosphatase, non-receptor type 2 (PTPN2), promyelocytic leukaemia protein (PML) or small ubiquitin-related modifier 1 (SUMO1), play a minor role in OSM-mediated induction of RLR. Remarkably, OSM and interferon-γ (IFN-γ) synergize to mediate transcription of RLR and pre-treatment of fibroblasts with OSM fosters the type I interferon production in response to a subsequent encounter with double-stranded RNA. Together, these findings suggest that the OSM-induced JAK/STAT1 signalling is implicated in virus protection of non-professional immune cells and may cooperate with interferons to enhance RLR expression in these cells. KW - oncostatin M KW - DExD/H-Box RNA helicase KW - RIG-I KW - STAT1 KW - innate immunity Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-159558 VL - 21 IS - 11 ER - TY - JOUR A1 - Schuster, Sarah A1 - Krüger, Timothy A1 - Subota, Ines A1 - Thusek, Sina A1 - Rotureau, Brice A1 - Beilhack, Andreas A1 - Engstler, Markus T1 - Developmental adaptations of trypanosome motility to the tsetse fly host environments unravel a multifaceted in vivo microswimmer system JF - eLife N2 - The highly motile and versatile protozoan pathogen Trypanosoma brucei undergoes a complex life cycle in the tsetse fly. Here we introduce the host insect as an expedient model environment for microswimmer research, as it allows examination of microbial motion within a diversified, secluded and yet microscopically tractable space. During their week-long journey through the different microenvironments of the fly´s interior organs, the incessantly swimming trypanosomes cross various barriers and confined surroundings, with concurrently occurring major changes of parasite cell architecture. Multicolour light sheet fluorescence microscopy provided information about tsetse tissue topology with unprecedented resolution and allowed the first 3D analysis of the infection process. High-speed fluorescence microscopy illuminated the versatile behaviour of trypanosome developmental stages, ranging from solitary motion and near-wall swimming to collective motility in synchronised swarms and in confinement. We correlate the microenvironments and trypanosome morphologies to high-speed motility data, which paves the way for cross-disciplinary microswimmer research in a naturally evolved environment. KW - none KW - tsetse fly KW - Trypanosoma KW - biophysics KW - microswimmer KW - sleeping sickness KW - structural biology Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158662 VL - 6 ER -