Immunisierung gegen das Tumorantigen CEA: Vergleichende Untersuchung an Wildtyp- und CEA transgenen Mäusen

Immunization protocols in animal models have shown that the tumor-associated antigen CEA could be a target for active immunization. Since human CEA which does not exist in the rodent genome has been used in these models it is hard to decide whether part of the immune response is directed only to the foreign antigen (anti-allo). We established an animal model on H2d background using CEA transfected tumor cells and CEA transgenic mice. Transplanting CEA transfected cells s.c. in wild type mice clearly shows, that the animals build up an immune response with high antibody titers in serum. Accordingly, only 55% of the animals developed solid tumors after tumor challenge. A cellular immune response could not be detected. In contrast, CEA transgenic mice did not develop any immune response and accepted the tumor in 90 % thus demonstrating the role of the foreign antigen. Active immunization using tumor lysates or lymphocytes loaded with human CEA was able to induce a CTL response as well as tumor-rejection in up to 76% of wild type mice. The immune response after intraperitoneal tumor lysate vaccination was not CEA specific and was able to lyse non-transfected CEA negative wild type tumor cells as well. CEA-transgenic mice could be induced to build up a CTL response in vitro with both vaccination protocols. In vivo CEA-transgenic mice rejected CEA-positive tumors in about 60% after immunization with the tumor lysate but not after intravenous vaccination with the CEA-loaded autologous lymphocytes. As there was a big difference in the reaction of the immune response to CEA between wild-type and transgenic rodents, the experiments clearly show the importance of transgenic models when testing the effects of immunization towards human tumor associated antigens. The induction of cross-priming with antigens other than CEA or the use of (co)stimulatory molecules must be taken into account when using CEA in vector systems for immunization.Die Immunisierung gegen ein Tumorantigen stellt eine Option in der Therapie von Tumoren, bzw. deren Metastasen dar. Während in Tiermodellen dabei über gute Erfolge berichtet wird, sind die klinischen Erfolge bis auf Einzelfälle eher entmutigend. Ein Grund für diese Diskrepanz könnte in der Verwendung ungeeigneter Modelle im Tierversuch liegen. So könnte die experimentelle Verwendung humaner Tumorantigene wie CEA in Nagern, in denen es kein identisches Gen gibt, zu einer immunologischen Reaktion des Tierorganismus führen. Um diese Frage zu überprüfen, wurde eine Tumorzelllinie mit dem häufigen Tumorantigen CEA transfiziert und das Wachstum dieser Zelllinie in Wildtypmäusen und einem CEA transgenen Tiermodell untersucht. An diesem Modell wurden dann unterschiedliche Methoden der Immunisierung getestet. Nach subkutaner Injektion einer Wildtypleukämiezelllinie und einer CEA transfizierten Leukämiezelllinie bei Wildtypmäusen des Stammes DBA/2N zeigte sich, dass 91% der nicht transfizierten Tumorzellen anwuchsen, während nur 55% der CEA transfizierten Tumorzellen eine Geschwulst bildeten. Eine Ursache für die niedrigere Anwachsrate der transfizierten Zellen liegt in der Erkennung des CEA durch das Immunsystem. So wurden Antikörper gegen CEA gebildet. Zytotoxische T-Lymphozyten konnten spontan nicht induziert werden. Wurden nun die Tumorzellen den CEA transgenen Mäusen injiziert, zeigte sich kein Unterschied in der Anwachsrate der Tumoren (90% Anwachsrate der transfizierten Zellen versus 90% Anwachsrate der Wildtypzellen). Folgerichtig konnten weder Antikörpertiter, noch zytotoxische T-Zellen gegen CEA nachgewiesen werden. An diesem Modell wurde dann eine Immunisierung mit Tumorzelllysat bestehend aus CEA transfizierten Zellen in kompletten Freundschen Adjuvans getestet. Damit konnte sowohl in den Wildtypmäusen, als auch in den transgenen Nagern eine in vitro und in vivo messbare zytotoxische Antwort gegen CEA exprimierende Zellen induziert werden. Allerdings war diese Antwort nicht CEA spezifisch, es wurde auch die Wildtypzelllinie abgestoßen. Damit ist gezeigt, dass die Immunantwort nach Vakzinierung mit Tumorzelllysat nicht nur CEA, sondern auch andere Antigene erkennt. Eine Möglichkeit, die CEA-Spezifität der Immunantwort weiter zu überprüfen, besteht in der intravenösen Immunisierung mit autologen Milzlymphozyten, die mit CEA beladen wurden. So konnte bei den syngenen Tieren eine Zytotoxizität gegen CEA exprimierende Zellen in vitro von bis zu 40% der Zielzellen festgestellt werden (E/T=50). Auch die transgenen Mäuse zeigten eine in vitro messbare Zytotoxizität von bis zu 20% der Zielzellen (E/T=10). In vivo konnte die Tumoranwachsrate der CEA positiven Tumorzellen in den Wildtyptieren von 55% auf 36% reduziert werden, während in den transgenen Tieren keine Reduktion nachgewiesen wurde. Gegen die CEA negative Mutterzelllinie zeigte sich keine immunologische Reaktion. Das Modell demonstrierte Unterschiede im immunologischen Verhalten der Wildtypmäuse und der transgenen Mäuse nach Implantation CEA exprimierender Tumorzellen. Im transgenen Modell, das der Situation im Menschen sehr nahe kommt, war im Gegensatz zum syngenen Modell keine spontane Immunantwort gegen CEA vorhanden. Die Vakzinierung mit einem Tumorzelllysat konnte in beiden Mäusestämmen eine effektive Immunantwort induzieren, die aber nicht nur gegen CEA, sondern auch gegen andere Antigene gerichtet ist. Eine spezifisch gegen CEA gerichtete Immunisierung mit beladenen Lymphozyten zeigte nur in den Wildtypmäusen einen in vivo messbaren Erfolg. Diese Beobachtungen zeigen die Bedeutung transgener Modelle bei der Durchführung von Vakzinierungsstudien gegen CEA und begründen teilweise die große Diskrepanz zwischen erfolgreichen Tierversuchen und den erfolglosen klinischen Studien, die bisher nur in einzelnen Patienten von einer in vivo wirksamen Immunisierung berichten. Sie demonstrieren auch, dass es mit entsprechenden Methoden, die mehrere Antigene bei der Vakzinierung benutzen, durchaus gelingen kann, die Toleranz gegen Tumorzellen zu durchbrechen und eine wirksame Immunantwort aufzubauen

Zeitschrift für Praktische Philosophie / Einleitung : Das Schöne, Wahre und Gute Das sinnvolle Leben in der Diskussion

Die Kategorie des Sinns oder des Sinnvollen findet in der jüngeren Diskussion um die normative Theorie des guten Lebens erhöhte Aufmerksamkeit. Viele Autoren gehen mittlerweile davon aus, dass eine umfassende Bestimmung des guten Lebens nicht mehr ohne die Erwähnung und Erläuterung der Wertdimension des sinnvollen Lebens auskommt. Typischerweise wird dieses Syntagma so verstanden, dass damit ein Wertaspekt des (bisherigen) Teil- oder Gesamtlebens eines (menschlichen) Individuums gemeint ist, welches in einer bestimmten, von anderen Wertdimensionen des guten Lebens noch abzugrenzenden Hinsicht als lobenswert, bedeutsam oder bewundernswert ausgezeichnet wird. Umstritten ist dabei jedoch zum einen, ob das sinnvolle Leben eine eigenständige Wertkategorie innerhalb des guten Lebens darstellt, und zum anderen, falls dem so wäre, wie diese Kategorie dann genau zu explizieren wäre, nicht zuletzt im Verhältnis zu den etablierten Wertkategorien innerhalb des guten Lebens, wie etwa dem Wohlergehen. Die in diesem Schwerpunkt versammelten Aufsätze unternehmen den Versuch, zum Verständnis der vielschichtigen und kontrovers diskutierten Dimension des Sinns beizutragen.The category of meaning or meaningfulness recently has received widespread attention within the debate on the good life. Indeed, many authors suppose that any comprehensive normative theory of the good life needs to include an account of the meaningful life, understood as one of its particular value dimensions. Usually, this syntagma is understood in terms of referring to a certain valuable aspect of a given life or one of its temporal parts, labelled as praiseworthy, significant, or admirable. Controversy abounds, however, when it comes to questions such as whether the meaningful life constitutes an independent value category within the good life, or, if so, how this category relates to the other, more established categories of the good life like human well-being. The articles collected in this special issue attempt to contribute to the controversies on the understanding of the multi-layered dimension of the meaningful life and try to shed further light on it.(VLID)325093

NRP2 transcriptionally regulates its downstream effector WDFY1.

Neuropilins (NRPs) are cell surface glycoproteins that often act as co-receptors for plexins and VEGF family receptors. Neuropilin-2 (NRP2), a family member of NRPs, was shown to regulate autophagy and endocytic trafficking in cancer cells, a function distinctly different from its role as a co-receptor. WD Repeat and FYVE domain containing 1 (WDFY1)-protein acts downstream of NRP2 for this function. Our results indicated that NRP2 maintains an optimum concentration of WDFY1 by negatively regulating its expression. Since increased expression of WDFY1 reduces the endocytic activity, maintenance of WDFY1 level is crucial in metastatic cancer cells to sustain high endocytic activity, essential for promotion of oncogenic activation and cancer cell survival. Here, we have delineated the underlying molecular mechanism of WDFY1 synthesis by NRP2. Our results indicated that NRP2 inhibits WDFY1 transcription by preventing the nuclear localization of a transcription factor, Fetal ALZ50-reactive clone 1 (FAC1). Our finding is novel as transcriptional regulation of a gene by NRP2 axis has not been reported previously. Regulation of WDFY1 transcription by NRP2 axis is a critical event in maintaining metastatic phenotype in cancer cells. Thus, inhibiting NRP2 or hyper-activating WDFY1 can be an effective strategy to induce cell death in metastatic cancer

The Role of lncRNAs TAPIR-1 and -2 as Diagnostic Markers and Potential Therapeutic Targets in Prostate Cancer

In search of new biomarkers suitable for the diagnosis and treatment of prostate cancer, genome-wide transcriptome sequencing was carried out with tissue specimens from 40 prostate cancer (PCa) and 8 benign prostate hyperplasia patients. We identified two intergenic long non-coding transcripts, located in close genomic proximity, which are highly expressed in PCa. Microarray studies on a larger cohort comprising 155 patients showed a profound diagnostic potential of these transcripts (AUC~0.94), which we designated as tumor associated prostate cancer increased lncRNA (TAPIR-1 and -2). To test their therapeutic potential, knockdown experiments with siRNA were carried out. The knockdown caused an increase in the p53/TP53 tumor suppressor protein level followed by downregulation of a large number of cell cycle- and DNA-damage repair key regulators. Furthermore, in radiation therapy resistant tumor cells, the knockdown leads to a renewed sensitization of these cells to radiation treatment. Accordingly, in a preclinical PCa xenograft model in mice, the systemic application of nanoparticles loaded with siRNA targeting TAPIR-1 significantly reduced tumor growth. These findings point to a crucial role of TAPIR-1 and -2 in PCa

An epigenetic reprogramming strategy to re-sensitize radioresistant prostate cancer cells

Radiotherapy is a mainstay of curative prostate cancer treatment, but risks of recurrence after treatment remain significant in locally advanced disease. Given that tumor relapse can be attributed to a population of cancer stem cells (CSC) that survives radiotherapy, analysis of this cell population might illuminate tactics to personalize treatment. However, this direction remains challenging given the plastic nature of prostate cancers following treatment. We show here that irradiating prostate cancer cells stimulates a durable upregulation of stem cell markers that epigenetically reprogram these cells. In both tumorigenic and radioresistant cell populations, a phenotypic switch occurred during a course of radiotherapy that was associated with stable genetic and epigenetic changes. Specifically, we found that irradiation triggered histone H3 methylation at the promoter of the CSC marker aldehyde dehydrogenase 1A1 (ALDH1A1), stimulating its gene transcription. Inhibiting this methylation event triggered apoptosis, promoted radiosensitization, and hindered tumorigenicity of radioresistant prostate cancer cells. Overall, our results suggest that epigenetic therapies may restore the cytotoxic effects of irradiation in radioresistant CSC populations

The empirical equilibrium structure of diacetylene

High-level quantum-chemical calculations are reported at the MP2 and CCSD(T) levels of theory for the equilibrium structure and the harmonic and anharmonic force fields of diacetylene, HCCCCH. The calculations were performed employing Dunning's hierarchy of correlation-consistent basis sets cc-pVXZ, cc-pCVXZ, and cc-pwCVXZ, as well as the ANO2 basis set of Almloef and Taylor. An empirical equilibrium structure based on experimental rotational constants for thirteen isotopic species of diacetylene and computed zero-point vibrational corrections is determined (r_e^emp: rC-H=1.0615 A, rCtripleC=1.2085 A, rC-C = 1.3727 A) and in good agreement with the best theoretical structure (CCSD(T)/cc-pCV5Z: rC-H=1.0617 Angstrom, rCtripleC=1.2083 A, rC-C=1.3737 A). In addition, the computed fundamental vibrational frequencies are compared with the available experimental data and found in satisfactory agreement.Comment: 12 pages, accepted for publication in J. Mol. Spectros

Neuropilin-2 regulates androgen-receptor transcriptional activity in advanced prostate cancer

Aberrant transcriptional activity of androgen receptor (AR) is one of the dominant mechanisms for developing of castration-resistant prostate cancer (CRPC). Analyzing AR-transcriptional complex related to CRPC is therefore important towards understanding the mechanism of therapy-resistance. While studying its mechanism, we observed that a transmembrane protein called neuropilin-2 (NRP2) plays a contributory role in forming a novel AR-transcriptional complex containing nuclear pore proteins. Using immunogold electron microscopy, high-resolution confocal microscopy, chromatin immunoprecipitation, proteomics, and other biochemical techniques, we delineated the molecular mechanism of how a specific splice variant of NRP2 becomes sumoylated upon ligand stimulation and translocates to the inner nuclear membrane. This splice variant of NRP2 then stabilizes the complex between AR and nuclear pore proteins to promote CRPC specific gene expression. Both full-length and splice variants of AR have been identified in this specific transcriptional complex. In vitro cell line-based assays indicated that depletion of NRP2 not only destabilizes the AR-nuclear pore protein interaction but also inhibits the transcriptional activities of AR. Using an in vivo bone metastasis model, we showed that the inhibition of NRP2 led to the sensitization of CRPC cells toward established anti-AR therapies such as enzalutamide. Overall, our finding emphasize the importance of combinatorial inhibition of NRP2 and AR as an effective therapeutic strategy against treatment refractory prostate cancer

German-wide prospective DACAPO cohort of survivors of the acute respiratory distress syndrome (ARDS): a cohort profile

Purpose While most research focuses on the association between medical characteristics and residual morbidity of survivors of the acute respiratory distress syndrome (ARDS), little is known about the relation between potentially modifiable intensive care unit (ICU) features and the course of health-related quality of life (HRQoL). Accordingly, the DACAPO study was set up to elucidate the influence of quality of intensive care on HRQoL and return to work (RtW) in survivors of ARDS. The continued follow-up of these former ICU patients leads to the establishment of the DACAPO (survivor) cohort. Participants Sixty-one ICUs all over Germany recruited patients with ARDS between September 2014 and April 2016. Inclusion criteria were: (1) age older than 18 years and (2) ARDS diagnosis according to the ‘Berlin definition’. No further inclusion or exclusion criteria were applied. 1225 patients with ARDS could be included in the DACAPO ICU sample. Subsequently, the 876 survivors at ICU discharge form the actual DACAPO cohort. Findings to date The recruitment of the participants of the DACAPO cohort and the baseline data collection has been completed. The care-related data of the DACAPO cohort reveal a high proportion of adverse events (in particular, hypoglycaemia and reintubation). However, evidence-based supportive measures were applied frequently. Future plans Three months, 6 months and 1 year after ICU admission a follow-up assessment is conducted. The instruments of the follow-up questionnaires comprise the domains: (A) HRQoL, (B) RtW, (C) general disability, (D) psychiatric symptoms and (E) social support. Additionally, an annual follow-up of the DACAPO cohort focusing on HRQoL, psychiatric symptoms and healthcare utilisation will be conducted. Furthermore, several add-on projects affecting medical issues are envisaged

German-wide prospective DACAPO cohort of survivors of the acute respiratory distress syndrome (ARDS): a cohort profile

Purpose While most research focuses on the association between medical characteristics and residual morbidity of survivors of the acute respiratory distress syndrome (ARDS), little is known about the relation between potentially modifiable intensive care unit (ICU) features and the course of health-related quality of life (HRQoL). Accordingly, the DACAPO study was set up to elucidate the influence of quality of intensive care on HRQoL and return to work (RtW) in survivors of ARDS. The continued follow-up of these former ICU patients leads to the establishment of the DACAPO (survivor) cohort. Participants Sixty-one ICUs all over Germany recruited patients with ARDS between September 2014 and April 2016. Inclusion criteria were: (1) age older than 18 years and (2) ARDS diagnosis according to the 'Berlin definition'. No further inclusion or exclusion criteria were applied. 1225 patients with ARDS could be included in the DACAPO ICU sample. Subsequently, the 876 survivors at ICU discharge form the actual DACAPO cohort. Findings to date The recruitment of the participants of the DACAPO cohort and the baseline data collection has been completed. The care-related data of the DACAPO cohort reveal a high proportion of adverse events (in particular, hypoglycaemia and reintubation). However, evidence-based supportive measures were applied frequently. Future plans Three months, 6 months and 1 year after ICU admission a follow-up assessment is conducted. The instruments of the follow-up questionnaires comprise the domains: (A) HRQoL, (B) RtW, (C) general disability, (D) psychiatric symptoms and (E) social support. Additionally, an annual follow-up of the DACAPO cohort focusing on HRQoL, psychiatric symptoms and healthcare utilisation will be conducted. Furthermore, several add-on projects affecting medical issues are envisaged

Characteristics and provision of care of patients with the acute respiratory distress syndrome: descriptive findings from the DACAPO cohort baseline and comparison with international findings: a cross-sectional study

Background: Little is known about the characteristics and real world life circumstances of ARDS (acute respiratory distress syndrome) patient populations. This knowledge is essential for transferring evidence-based therapy into routine healthcare. The aim of this study was to report socio-demographic and clinical characteristics in an unselected population of ARDS patients and to compare these results to findings from other large ARDS cohorts. Methods: A German based cross-sectional observational study was carried out. A total of 700 ARDS patients were recruited in 59 study sites between September 2014 and January 2016. Socio-demographic, disease and care related variables were recorded. Additionally, characteristics of other large ARDS cohorts identified by a systematic literature search were extracted into evidence tables. Results: Median age of ARDS patients was 58 years, 69% were male. Sixty percent had no employment, predominantly due to retirement. Seventy-one percent lived with a partner. The main cause of ARDS was a pulmonary 'direct' origin (79%). The distribution of severity was as follows: mild (14%), moderate (48%), severe (38%). Overall ICU mortality was calculated to be 34%. The observed prevalence of critical events (hypoxemia, hypoglycemia, re-intubation) was 47%. Supportive measures during ICU-treatment were applied to 60% of the patients. Other ARDS cohorts revealed a high heterogeneity in reported concomitant diseases, but sepsis and pneumonia were most frequently reported. Mean age ranged from 54 to 71 years and most patients were male. Other socio-demographic factors have been almost neglected. Conclusions: The proportion of patients suffering of mild ARDS was lower compared to the only study identified, which also applied the Berlin definition. The frequency of critical events during ICU treatment was high and the implementation of evidence-based therapy (prone positioning, neuro-muscular blockers) was limited. More evidence on socio-demographic characteristics and further studies applying the current diagnostic criteria are desirable