Verlauf der Knochendichte und der Knochenumbaumarker unter der Therapie mit Romosozumab

As a result of demographic changes and the associated increase in life expectancy, pharmacological treatment of osteoporosis will continue to gain importance in the coming years. Currently, the monoclonal antibody romosozumab represents the most effective form of therapy for women with postmenopausal osteoporosis. In the two pivotal approval studies (FRAME and ARCH), increases in bone mineral density (BMD) at the lumbar spine by 13,3% and 13,7%, and at the total hip by 6,8% and 6,2%, respectively, were demonstrated. In this study, data from 68 female patients who were treated with romosozumab for postmenopausal osteoporosis at the Endokrinologikum Göttingen were retrospectively analyzed. A significant increase in BMD was observed at the lumbar spine by 22,2% and at the total left hip by 11,1% (p<0,001). Thus, this real-world clinical study showed a greater increase in BMD than the approval studies. When comparing the patient populations, it became apparent that our patients had the lowest baseline BMD at the lumbar spine (T-score: -3,6 SD) and, along with the ARCH patients, the lowest BMD at the total left hip (T-score: -2,8 SD) before therapy. Additionally, our patients were 6,2 years younger than those in the FRAME study and had more vertebral fractures (60,3% vs. 18,7%), suggesting a sicker cohort compared to the FRAME study. Compared to the ARCH study, our patients were also 9,7 years younger but had fewer vertebral fractures (ARCH study: 96,2%). While the participants in the approval studies did not receive prior treatment, 69,1% of the patients in our study had undergone prior therapy for 4,8 years. Among them, patients with previous teriparatide treatment had the greatest BMD increase, while those previously treated with denosumab showed the least improvement. We found that patients experienced a similar BMD increase regardless of the presence of secondary risk factors. Furthermore, it was observed that baseline BAP concentrations correlated with the BMD increase at the total left hip (r=0,4; p<0,05). Patients in our study also had a higher calcium intake than those in the approval studies. It is therefore conceivable that the higher baseline BAP levels and increased calcium intake contributed to the greater BMD increase. Additionally, the younger age of our patients may have played a role in this improved outcome. Analysis of bone turnover markers revealed a significant increase in the bone formation markers P1NP (p<0,01), osteocalcin (p<0,05), and BAP (p<0,001) after one month of therapy. These markers fell below baseline levels after six to nine months, clearly demonstrating the initial osteoanabolic effect of romosozumab. The antiresorptive effect of romosozumab was reflected by the course of the bone resorption marker CTX, whose concentration decreased after initiation of romosozumab therapy. Moreover, baseline levels of remodeling markers provided insights into the type of prior treatment. During romosozumab therapy, only one patient sustained a traumatic—i.e., non-osteoporotic—fracture.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt. !Es liegt auch ein Embargo vor!These files are not accessible until the state exam has been completed

In-ovo studies on the cellular metabolism of Hodgkin's lymphoma

Die ausgezeichneten Überlebenschancen von Patienten mit Hodgkin-Lymphom (HL), einer bösartigen Neoplasie des lymphatischen Systems, können durch akute und langfristige Toxizitäten wie Infertilität, kardiovaskuläre Schäden oder sekundäre Malignome beeinträchtigt werden. Darüber hinaus werden dringend neue Therapien für Patient*innen benötigt, die nach einer Hochdosis-Chemotherapie einen Rückfall erleiden, mit dem Ziel, unerwünschte Risiken zu minimieren und gleichzeitig die Wirksamkeit zu erhalten. Zu den aufkommenden Therapieansätzen gehören kleine Moleküle, die intrazelluläre Proteine gezielt angreifen. Ein Beispiel dafür ist das Enzym Nicotinamidphosphoribosyltransferase (NAMPT), das in verschiedenen Tumorentitäten aberrant aktiv ist. In-vitro-Analysen haben gezeigt, dass chemische Inhibitoren der NAMPT toxisch für Zellen des HL sein können. Deshalb ist davon auszugehen, dass die NAMPT als geschwindigkeitsbestimmendes Enzym des Nicotinamidadenindinukleotid (NAD⁺)-Stoffwechsels wichtig für das HL ist. Es ist jedoch noch unklar, ob eine Intervention der NAMPT-Aktivität auch in vivo das Lymphomwachstum von HL-Zellen hemmt. Ziel der vorliegenden Studie war es deshalb, ein präklinisches Xenograft-Modell zu nutzen. In dem In-ovo-Xenograft-Modell der Chorioallantoismembran des befruchteten Hühnereis (CAM) sollte untersucht werden, inwieweit die Inhibitoren FK866 und OT-82 die Lymphomentwicklung der L428-HL-Zellen verändern, sowie eine rot fluoreszierende, mCherry exprimierende Zelllinie für weiterführende In-ovo-Metabolismus-Analysen zu etablieren. Es wurden Zellen der HL-Zelllinie L428 auf die CAM appliziert, die zuvor 24 Stunden mit FK866, OT-82 oder zur Kontrolle mit DMSO behandelt wurden. Die entstandenen Läsionen wurden nach 96 Stunden geerntet und histologisch sowie morphologisch charakterisiert. Zusätzlich zur Oberfläche wurde das Volumen der Tumore mittels Mikro-Computertomographie (micro-CT) bestimmt. Die mit FK866 und OT-82 behandelten Zellen waren noch in der Lage, in ovo Lymphome zu bilden, sie waren aber kleiner, weniger durchblutet und von geringerer Zelldichte bei höherem Kollagengehalt. Die Tumorzellen waren allerdings positiv für den Proliferationsmarker Ki67. Somit führt die Behandlung mit NAMPT-Inhibitoren zu einer Reduzierung von Tumorgröße und -durchblutung, nicht jedoch zu einem vollständigen therapeutischen Effekt. Um zukünftige metabolische Analysen in ovo durchführen zu können, wurde eine Zelllinie etabliert, die das rot fluoreszierende Fusionsprotein mCherry-H2b exprimiert. Die hierbei etablierten Zelllinien reagierten in In-vitro- sowie in In-ovo-Studien ähnlich wie die parentale Zelllinie L428 auf die Behandlung mit den NAMPT-Inhibitoren FK866 und OT-82 hinsichtlich Zellviabilität, NAD+-Gehalt und NAMPT-Expression. Dies weist darauf hin, dass die etablierten Zelllinien für zukünftige Metabolismus-Analysen geeignet sind. Zusammengefasst konnte ein Xenograft-Modell etablieren, das zur Analyse der NAMPT-Aktivität und deren Einfluss auf die Lymphomentwicklung geeignet ist. Weiterhin konnten wir zeigen, dass die NAMPT-Inhibition Einfluss auf die Lymphomentwicklung in ovo haben. Somit steht ein neues präklinisches Modell zur Untersuchung des Lymphommetabolismus, beispielsweise durch Fluoreszenzlebensdauer-Mikroskopie, zur Verfügung.Excellent survival rates of patients with Hodgkin lymphoma (HL), a malignant neoplasm of the lymphatic system, may be compromised by acute and long-term toxicities, including infertility, cardiovascular damage, and secondary malignancies. Furthermore, there is an urgent need for novel therapeutic strategies for patients who experience relapse following high-dose chemotherapy, with the aim of minimizing adverse effects while maintaining therapeutic efficacy. Upcoming treatment approaches include small molecules that selectively target intracellular proteins. One such target is the enzyme nicotinamide phosphoribosyltransferase (NAMPT), which has been found to be aberrantly active in various tumor entities. In vitro analyses have shown that chemical inhibition of NAMPT can be cytotoxic to HL cells. Consequently, as a rate-limiting enzyme in the nicotinamideadeninedinucleotide (NAD⁺) metabolism, NAMPT is presumed to play a crucial role in HL cells. However, it remains unclear whether inhibiting NAMPT activity can also suppress HL tumor growth in vivo. The present study aimed to establish a preclinical xenograft model to investigate this question. Specifically, an in-ovo xenograft model of the chorio-allantoic membrane (CAM) of fertilized chicken eggs was employed to evaluate the effects of the NAMPT inhibitors FK866 and OT-82 on the development of L428 HL cells. This model has been shown to reflect key aspects of human lymphoma progression. Additionally, a red fluorescent mCherry-expressing cell line was established for future in-ovo metabolic analyses. Cells of the L428 HL cell line were applied to the CAM following a 24-hour pre-treatment with FK866, OT-82, or dimethyl sulfoxide (DMSO) as a control. After 96 hours, the resulting lesions were harvested and characterized histologically and morphologically. In addition to tumor surface area, tumor volume was quantified using micro-computed tomography (micro-CT). Although FK866- and OT-82-treated HL cells were still capable of forming tumors in ovo, the resulting lymphomas were smaller, less vascularized, and exhibited lower cellular density with an increased collagen content. However, tumor cells remained positive for the proliferation marker Ki67. Thus, NAMPT inhibition led to a reduction in tumor size and vascularization but did not achieve a complete therapeutic effect. To facilitate future metabolic analyses in ovo, a cell line expressing the red fluorescent fusion protein mCherry-H2B was established. This newly generated cell line responded similarly to the parental L428 cell line in both in-vitro and in-ovo studies regarding cell viability, NAD⁺ levels, and NAMPT expression after treatment with FK866 and OT-82. This suggests that the established cell lines are suitable for future metabolism-focused investigations. In summary, we successfully developed a xenograft model that enables the study of NAMPT activity and its impact on HL tumor development. Furthermore, we demonstrated that NAMPT inhibition influences lymphoma progression in ovo. This novel preclinical model provides a valuable tool for investigating lymphoma metabolism, including fluorescence lifetime microscopy-based analyses.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt. !Es liegt auch ein Embargo vor!These files are not accessible until the state exam has been completed

Establishment and characterization of cervical carcinoma cell lines with stable overexpression of the G-protein-coupled estrogen receptor (GPER1)

Cervical cancer (CC) remains the fourth most common malignancy among women worldwide, despite effective HPV vaccination strategies. CC can be classified into different histological subtypes. The majority of cases are cervical squamous cell carcinomas (CSCC), followed by adenocarcinomas (CAC), the latter including a higher proportion of HPV-negative tumors. These subtypes differ in their etiology, pathology, and clinical presentation. While the incidence of CSCC has been declining, CAC incidence has increased in recent years. GPER1 is a membrane-bound G protein-coupled receptor with seven transmembrane domains that activates various intracellular signaling cascades. These include cAMP production, the Rho-ROCK pathway, and EGFR transactivation, which subsequently stimulate MAPK and Akt pathways. Through HIF-1α, GPER1 further modulates NOTCH and VEGF signaling. GPER1 plays a relevant role in the initiation and progression of several malignancies, including CC. To elucidate the specific role of GPER1 in CC, stable overexpression (OE) of GPER1 was established in the CC cell lines SiHa and HeLa using a non-viral Sleeping-Beauty transposon-based vector. Stable GPER1-OE was confirmed by RT-qPCR, Western blotting, and flow cytometry (FACS). Functional consequences of GPER1-OE were assessed by examining tumor-associated cellular behaviors including proliferation, migration, invasion, apoptosis, metabolic viability, as well as stem-cell-associated properties via colony- and tumorsphere-formation assays. In addition, next-generation sequencing (NGS) was conducted to identify differential gene expression and signaling pathways particularly relevant for tumor development and progression. Overall, GPER1-OE resulted in a more aggressive phenotype in SiHa cells (CSCC), characterized by increased proliferation, migration, and enhanced cancer stem-cell-like traits. In contrast, HeLa cells (CAC) displayed a less aggressive phenotype, with reduced proliferation and migration, increased apoptosis, and diminished stem-cell properties. The NGS analysis supported these findings, revealing an upregulation of mTOR, MYC, p53, EMT, and angiogenesis-related pathways in SiHa cells with GPER1-OE, whereas these pathways were predominantly downregulated in HeLa cells, accompanied by suppressed Hedgehog, KRAS, and Wnt/β-catenin signaling. These results suggest that GPER1 can exert either tumor-promoting or tumor-suppressive effects in CC depending on the histological subtype.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt.These files are not accessible until the state exam has been completed

Schmidt-Lanterman incisures in the Schwann cell and their role for peripheral nerve regeneration

Schmidt-Lanterman incisures (SLIs) are cytoplasmic channels within compact peripheral myelin, yet their formation and functional relevance in health and disease remain incompletely understood. This study investigates SLI development, their regulation across the lifespan, and their role in neuropathy and nerve regeneration. Using immunofluorescence and high-pressure freezing electron microscopy of developing murine sciatic nerves, we show that SLIs arise early during myelin formation. During development, SLI number increases in parallel with internodal elongation and remains stable in adulthood and aging, indicating an essential role in Schwann cell maturation and myelin maintenance. In mouse models of PMP22-associated neuropathies (CMT1A and HNPP), SLI number is altered in a dosage-dependent manner: increased in PMP22-duplication and transiently reduced during early development in PMP22-deletion. Similarly, sural nerve biopsies from patients with demyelinating neuropathy reveal increased SLI numbers, particularly in short internodes, suggesting a compensatory function in disease. Following sciatic nerve crush injury, mice with reduced SLI numbers (VCLcKO) exhibit delayed motor recovery and fail to upregulate SLIs, in contrast to wildtype controls. These findings support a critical role of SLIs in preserving nerve function and promoting regeneration, potentially by facilitating axo-glial communication and metabolic support. Together, this work identifies SLIs as dynamic and regulated structures essential for peripheral nerve development, maintenance, and repair, highlighting their potential as targets for therapeutic strategies to enhance nerve regeneration.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt.These files are not accessible until the state exam has been completed

Mode of action of direct KRAS inhibition and development of KRAS inhibitor-based combination therapies

As pancreatic ductal adenocarcinoma (PDAC) has still one of the least favourable survival rates among cancers, better treatment is urgently needed. This thesis aimed to make use of a new class of targeted therapies, the Kirsten Rat Sarcoma virus (KRAS)G12C inhibitors and to evaluate their selectivity and mechanisms in context of PDAC cell lines. Because these inhibitors are also known to be susceptible to resistance development, potential combination therapies were sought and evaluated. First of all, different KRASG12C inhibitors were tested on various PDAC cell lines with and without the KRASG12C mutation. Sotorasib, Adagrasib and MRTX-1257 revealed a selective reduction of cell viability in both Clonogenic and CellTiter-Glo® Assays specifically in KRASG12C mutated MiaPaCa2 cells. By the testing, Sotorasib was prioritized for the following experiments due to the specificity and the authorization for clinical use before this work has started. Further experiments were mainly done on human 2D cell lines which were either commercially available (MiaPaCa2) or generated from patient-derived organoids (PAN51T-2D). To comprehend the impact of KRASG12C inhibitors on signalling pathways, Western blot analysis was performed on cell lysates obtained from MiaPaCa2 and PAN51T-2D cells following different treatment durations of Sotorasib. On-target activity of Sotorasib was detected in both cell lines after all time points. In addition, a downregulation of downstream targets of KRAS was observed after a treatment time of 24 hours. However, after incubation with Sotorasib for 48 hours and 72 hours, a dose-dependent recovery of ERK phosphorylation was exhibited, suggesting that downstream pathways were reactivated, besides KRAS blockade. To overcome these resistance mechanisms, a Sotorasib-anchored unbiased drug screen with 126 different drugs, ranging from clinical approved drugs to preclinical compounds, was performed on MiaPaCa2 cells. Nine potential candidates were identified, of which three could be validated by Clonogenic, CellTiter-Glo® Assays, and the calculation of a Bliss-synergy score. These were the SHP2 inhibitor TNO155, the Son of Sevenless KRAS interaction inhibitor BI-3406, and the tyrosine kinase inhibitor Nintedanib. Ultimately, Western blot analysis was performed once more after treating both cell lines with Sotorasib and TNO155 in combination for 72 hours. This time, the pERK recovery could be prevented in the PAN51T-2D cell line, while the combined treatment had only little effect on the MiaPaCa2 cell line. Additionally, cell cycle analysis was performed via fluorescence-activated cell sorting, which found that a combination treatment led to an increased fraction of MiaPaCa2 cells entering G1 cell cycle arrest. Contrary, the treatment of PAN51T-2D cells with Sotorasib and TNO155 did not lead to a G1 cell cycle arrest. In summary, this work has identified the SHP2 inhibitor TNO155 as a potential synergistic partner for Sotorasib, with the ability to overcome potential resistance mechanisms. Nevertheless, more research is needed to fully understand the divergent cellular responses after the combination treatment.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt.These files are not accessible until the state exam has been completed

Differential diagnostic distinction between Panayiotopoulos syndrome and migraine

Migräne zählt zu den häufigsten primären Kopfschmerzsyndromen im Kindes- und Jugendalter. Das Panayiotopoulos-Syndrom (PS) (auch bekannt als early-onset benign occipital seizure susceptibility syndrome (EBOSS) oder self-limited epilepsy with autonomic seizures (SeLEAS)), eine gutartige fokale Epilepsie mit autonomem Symptombild, kann aufgrund seiner klinischen Präsentation leicht mit einer Migräne verwechselt werden. Die korrekte Unterscheidung ist jedoch essenziell, da sie eine adäquate, frühzeitige Behandlung nach sich ziehen und der Leidensdruck der Patient*innen und Eltern senken würde. Ziel dieser retrospektiven Studie war es, die Häufigkeit von PS bei Kindern und Jugendlichen zu untersuchen, die mit der Verdachtsdiagnose „Migräne“ oder „Kopfschmerzen“ in der neuropädiatrischen Sprechstunde der Universitätsmedizin Göttingen vorgestellt wurden. Zudem sollten die von Patient*innen und Eltern beschriebenen Symptome miteinander verglichen werden, um potenziell eine bessere Abgrenzung der beiden Krankheitsbilder anhand der Semiologie zu ermöglichen. Insgesamt wurden 186 Patient*innen im Alter von einem bis 14 Jahren anhand von Arztbriefen und EEG-Befunden ausgewertet. Anhand der Anamnese und der EEG-Befunde konnte das PS bei 16 (8,6%) der Kinder und Jugendlichen identifiziert werden, bei 12 dieser Patient*innen wurde die Diagnose zusätzlich durch eine deutliche Besserung unter antikonvulsiver Therapie bestätigt. Bei weiteren 19 (10,2%) Kindern und Jugendlichen wies das EEG Epilepsietypische Potentiale (ETPs) auf, die ebenfalls mit einem PS vereinbar wären. Das gehäufte Auftreten von Anfällen aus dem Schlaf sowie das gleichzeitige Vorhandensein von Übelkeit, Würgereiz und Erbrechen bei PS waren Merkmale, die signifikant seltener bei Migräne als bei dem PS beobachtet wurden. Die Ergebnisse zeigen, dass bei der diagnostischen Abklärung von Kindern und Jugendlichen mit Kopfschmerzen oder Migräne die wichtige Differenzialdiagnose des PS stärker als bisher berücksichtigt werden sollte. Um dieses Krankheitsbild nicht zu übersehen, sollte die diagnostische Abklärung eine gezielte Anamnese mit Schwerpunkt auf autonomen Begleitsymptomen sowie die Durchführung eines EEGs umfassen.Migraine is among the most common primary headache syndromes in childhood and adolescence. Panayiotopoulos syndrome (PS) (also known as early-onset benign occipital seizure susceptibility syndrome (EBOSS) or self-limited epilepsy with autonomic seizures (SeLEAS)), is a benign focal epilepsy characterized by autonomic symptoms. Due to its clinical presentation, PS can easily be mistaken for migraine. Correct differential diagnosis is essential, as it enables timely and appropriate treatment and can significantly reduce the burden on both patients and their families. The aim of this retrospective study was to investigate the frequency of PS among children and adolescents who were referred to the pediatric neurology outpatient clinic at the University Medical Center Göttingen with a suspected diagnosis of “migraine” or “headache.” In addition, the symptoms described by patients and their parents were systematically compared to identify possible semiological differences that may assist in distinguishing between PS and migraine. A total of 186 patients aged 1 to 14 years were included in the analysis, based on physician‘s letters and EEG findings. PS was diagnosed in 16 children (8.6%) based on clinical presentation and EEG results; in 12 of these cases, the diagnosis was further confirmed by a marked improvement under anti-seizure medication. An additional 19 patients (10.2%) showed EEG abnormalities consistent with PS. The frequent occurrence of seizures originating from sleep, along with the simultaneous presence of nausea, retching, and vomiting, were features observed significantly less often in migraine than in Panayiotopoulos syndrome. These findings highlight the need for greater attention to Panayiotopoulos syndrome (PS) as a differential diagnosis in the evaluation of headache in children and adolescents, compared to current clinical practice. To avoid overlooking this condition, the diagnostic work-up should include a targeted medical history focusing on the characteristic features of autonomic seizures (such as headache episodes arising from sleep, ictal gaze deviation, nausea, retching, vomiting, and impaired consciousness) as well as an EEG examination.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt.These files are not accessible until the state exam has been completed

Pankreatisch zystische Neoplasien und das orointestinale Mikrobiom: Evaluierung als Biomarker

Pancreatic cystic neoplasms (PCN) are mostly benign cystic lesions of the pancreas, which can be a precursor lesion for pancreatic cancer. The risk of malignancy varies widely, depending on the entity of the PCN. Despite known risk factors and several studies, it has not yet been possible to reliably predict which patients with PCN will develop worrisome features or high-grade dysplasia and will therefore benefit from an early operation. Currently, it was revealed the benign pancreatic cysts and the pancreatic cancer harbors an own microbiome, Furthermore, the orointestinal microbiome can be exploited as a predictor for a pancreatic mass dignity. However, there is still a lack of knowledge whether the microbiome can predict the course of PCN. The aim of this prospective study was to investigate the orointestinal microbiome of patients with PCN and the association with the cyst entities, the Dutch-American risk stratification tool (DART-1) score and the presence of worrisome features in branch-duct intraductal papillary mucinous neoplasm (BD-IPMN). Buccal and rectal swab samples were prospectively collected to analyze the microbiome of patients with PCN. The microbiome was assessed by Oxford Nanopore sequencing technology (16S rRNA and metagenomics). Clinical data was collected with a standardized online questionnaire (RedCap). 78 patients have been enrolled in the study between January 2021 and December 2022. Alpha diversity metrics as the Observed Species, Shannon Index and Inverse-Simpson Index were determined and compared between the patients. Furthermore, the relative difference between these groups was shown and calculated using beta diversity distance metrics. The buccal microbiome of patients with BD-IPMN with worrisome features, as defined by the Fukuoka guidelines of 2017, revealed to harbor significantly different species compared to those without worrisome features. The rectal microbiome showed no significant differences between the two groups. In addition, a higher DART-1 score in patients with BD-IPMN was associated with significantly lower alpha diversity. No significant differences in the buccal or rectal microbiome were found between the cyst entities. The present study provides an opportunity to further research the microbiome as a biomarker for the progression of PCN.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt.These files are not accessible until the state exam has been completed

The influence of histone H3 modifications on darolutamide resistance in prostate cancer

Prostate cancer is one of the most frequently diagnosed malignancies in men worldwide and accounts for a substantial proportion of cancer-related deaths. With the introduction of darolutamide, a next-generation androgen receptor (AR) antagonist, a novel therapeutic option with promising clinical results became available in 2020. However, with the expanded clinical use of darolutamide, an increasing number of resistant cases has been observed. Previous studies have identified epigenetic alterations as key drivers in the development and progression of prostate cancer. This study investigated the role of histone H3 lysine 27 trimethylation (H3K27me3) and its writer protein, Enhancer of Zeste Homolog 2 (EZH2), in the development of resistance to darolutamide. VCaP cells were used as an in vitro model of castration-resistant prostate cancer (CRPC) and were treated with increasing concentrations of darolutamide to induce resistance. Western blot analyses revealed elevated protein levels of the repressive histone modification H3K27me3 and EZH2 in darolutamide-resistant VCaP cells (VCaP Daro). These changes correlated with reduced transcription of H3K27me3 target genes, including insulin-like growth factor-binding protein 3 (IGFBP3), homeobox D1 (HOXD1), AR splice variant V7 (ARV7), and AR full-length (Arfl). EZH2 knockdown resulted in decreased H3K27me3 levels and increased transcription of target genes. Similar effects were observed after treatment with EPZ-6438, a selective inhibitor of EZH2 methyltransferase activity. Chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) revealed a global, genome-wide increase in H3K27me3 occupancy in VCaP Daro, accompanied by a redistribution of H3K27me3 peaks toward gene promoter regions, potentially associated with altered AR binding. Subsequent RNA sequencing identified the sonic hedgehog (SHH) signaling pathway as the most strongly upregulated pathway in VCaP Daro, suggesting a central role in the resistance mechanism. Increased transcription of GLI Family Zinc Finger proteins 1 and 2 (GLI1, GLI2) served as readouts for Hedgehog pathway activation. Hedgehog reporter assays demonstrated SHH ligand secretion in VCaP Daro but not in wild-type VCaP cells (VCaP Wt), indicating possible autocrine or paracrine activation. Furthermore, treatment with the Smoothened inhibitor vismodegib demonstrated that Hedgehog signaling contributes to increased EZH2 expression. Both EPZ-6438 and vismodegib significantly reduced cell viability in both cell lines, as determined by MTS assays. Notably, the inhibitory effects were significantly stronger in VCaP Daro than in VCaP Wt, with vismodegib exhibiting greater cytotoxicity than EPZ-6438. In conclusion, these results suggest that EZH2 and SHH represent promising therapeutic targets for the treatment of darolutamide-resistant CRPC. As corresponding inhibitors are already clinically available, there is substantial translational potential. Further studies are required to elucidate the underlying molecular mechanisms.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt.These files are not accessible until the state exam has been completed

Integration of biological characteristics of pancreatic cancer PDX and CDX models with annotations of the donor patients

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with a five-year survival rate of approximately 12%, underscoring the urgent need for improved therapeutic strategies. A deeper understanding of PDAC biology is essential and relies on preclinical model systems such as patient-derived xenografts (PDX) and cell-derived xenografts (CDX). However, the extent to which these models reflect the primary tumors and the respective donor patients’ disease course remains elusive. In this study, we characterized clinical, histological, and genomic data from the KFO5002 cohort, including 119 patients, 71 primary tumors, 43 PDX models, and 8 CDX models. All data was integrated and analyzed using a private space of the open source bioinformatical platform cBioPortal. Genomic comparisons revealed that PDX models showed minimal de novo mutations, while CDX models largely preserved the mutational profiles of their corresponding PDX tumors. Analysis of PDX engraftment demonstrated accelerated tumor growth across increasing mouse generations. The grading of primary and PDX tumors showed a positive correlation between primary tumors and first-generation PDX models and remained stable across subsequent generations. Immunohistochemical analyses indicated higher proliferative capacity in most PDX tumors compared to primary tumors and suggested a predominant classical PDAC subtype in most of the PDX tumors. CDX models displayed heterogeneous morphology, mutational landscapes, and proliferative capacities. Drug response assays using gemcitabine, 5-fluorouracil, and paclitaxel revealed variable treatment sensitivities. Although no significant correlation between CDX drug response and patient survival was observed, clinical survival data aligned with established treatment guidelines, highlighting the relevance of the KFO5002 data set. Overall, PDX and CDX models represent valuable but limited tools for PDAC research. Their complementary use, alongside emerging approaches such as patient-derived organoids, may better capture tumor heterogeneity and therapeutic response. This study further demonstrates the utility of cBioPortal as a suitable platform for integrated translational cancer research.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt.These files are not accessible until the state exam has been completed

Reevaluation der standardisierten CCT - Kontrollbildgebung bei Kindern nach neurochirurgischer Tumorresektion

Nach operativen Eingriffen im Bereich der Neurochirurgie wurde standardmäßig eine postoperative kraniale Bildgebung mittels Computertomographie (CCT) als angewandtes Kontrollverfahren herangezogen. Ziel war es, eine durch die Operation entstandene und therapeutisch relevante Komplikation zu detektieren oder auszuschließen. Jedoch birgt die routinemäßige Nutzung dieser postoperativen Bildgebung einige Nachteile, wie die Risiken durch die Strahlenbelastung, den intrahospitalen Transport und die entstehenden Kosten. Kinder gelten als besonders vulnerabel hinsichtlich diagnostischer Strahlenbelastung, und es besteht ein erwiesenermaßen signifikant höheres Risiko der Entstehung einer Neoplasie. Es galt, erstmalig zu evaluieren, inwieweit das frühpostoperative Kontroll-CCT, auch EPOCT genannt (early postoperative computed tomography), nach speziell pädiatrisch neurochirurgischer Tumorresektion für die postoperative Überwachung und Behandlung einen relevanten therapeutischen Nutzen hat. Innerhalb dieser retrospektiven klinischen Studie wurden Daten aller pädiatrischen Patienten des Schwerpunktes Kinderneurochirurgie der UMG untersucht, die sich zwischen 2011 und 08/2022 einer intrakraniellen Tumorresektion sowie innerhalb von vier Stunden einer EPOCT unterzogen haben. Es wurden selektierte Variablen gesammelt und hinsichtlich ihrer Assoziation mit dem Outcome der EPOCT einzeln untersucht. Im Anschluss wurden acht logistische Regressionsanalysen und ROC-Kurven erstellt, um mehrere Variablen gleichzeitig im gemeinsamen Einfluss auf das Ergebnis der Kontrollbildgebung darzustellen und eine Schätzung des Outcomes zu ermöglichen. Es erwiesen sich bestimmte Variablen als signifikant in der Assoziation mit einer auffälligen EPOCT: Die neurologischen Defizite unmittelbar postoperativ, die präoperativen Maße der Ventrikelweite ER und FOHR, eine präoperativ gelegte extraventrikuläre Drainage EVD, die OP-Dauer und eine intra- oder unmittelbar postoperative Erythrozytentransfusion. Im Rahmen der logistischen Regressionsanalysen konnten zur Schätzung des Outcomes der EPOCT drei der acht Modelle mit der höchsten Güte herausgestellt werden: Neurologische Defizite in Modell 1 mit EVD (AUC 0,82), in Modell 3 mit Tumorlokalisation (AUC 0,81) und in Modell 4 mit OP-Dauer (AUC 0,85). Es ließ sich jeweils ein Score erstellen, der eine Indikationsstellung für eine EPOCT erlaubt. Es zeigte sich, dass die neurologische Auffälligkeit einen starken Zusammenhang mit dem Ergebnis der EPOCT aufweist. In Anbetracht einer möglichen Indikation zur notfallmäßigen Rückkehr in den Operationssaal konnte gezeigt werden, dass die neurologische Untersuchung einen höheren Stellenwert einnahm als die EPOCT. In keinem einzigen Fall wurde eine notfallmäßige Reoperation allein durch eine auffällige Bildgebung indiziert. Zusammenfassend lässt sich festhalten, dass die routinemäßige EPOCT zur Indikationsstellung einer notfallmäßigen Reoperation nach neurochirurgischer Tumorresektion der klassischen neurologischen Untersuchung nicht überlegen ist. Es konnten Risikofaktoren identifiziert werden, die nachweislich signifikant mit einer auffälligen Kontrollbildgebung oder einer Veränderung im Patientenmanagement einhergingen. Zudem konnten Scores erstellt werden, die eine Einschätzung des Outcomes der EPOCT in kausalem Zusammenhang mit zwei Variablen ermöglichen. Nach aktuellem Stand ist es noch nicht möglich, einen exakten Score zu erstellen, der fallübergreifend als Wegweiser dienen kann. Bis dies gelingt, möglichst mittels einer größeren, prospektiven Studie, gilt es, die EPOCT von ihrer Position als routinemäßige Kontrolle abzulösen und sie lediglich als unterstützendes Hilfsmittel heranzuziehen.After surgical interventions in the field of neurosurgery, postoperative cranial imaging using computed tomography (CCT) was used as a standard control procedure. The aim was to detect or rule out any therapeutically relevant complications resulting from the operation. However, the routine use of this postoperative imaging has a number of disadvantages, such as the risks associated with radiation exposure, intrahospital transportation and the costs incurred. Children are considered to be particularly vulnerable to diagnostic radiation exposure, and there is a proven significantly higher risk of neoplasia developing. The aim was to evaluate for the first time the extent to which early postoperative computed tomography (EPOCT) has a relevant therapeutic benefit for postoperative monitoring and treatment after tumor resection specifically in pediatric neurosurgery. Within this retrospective clinical study, data of all pediatric patients of the department of pediatric neurosurgery of the UMG who underwent intracranial tumor resection and an EPOCT within four hours between 2011 and 08/2022 were examined. Selected variables were collected and individually inspected with regard to their association with the outcome of the EPOCT. Subsequently, eight logistic regression analyses and ROC curves were created to show the joint influence of several variables on the result of the control imaging and to enable an estimation of the outcome. Certain variables were found to be significantly associated with an abnormal outcome of the EPOCT: immediate postoperative neurologic deficits, preoperative measures of ventricular width ER and FOHR, a preoperatively placed extraventricular drain EVD, duration of surgery, and an intraoperative or immediate postoperative erythrocyte transfusion. Within the logistic regression analyses, three of the eight models with the highest quality were identified for estimating the outcome of the EPOCT: Neurological deficits in model 1 with EVD (AUC 0.82), in model 3 with tumor localisation (AUC 0.81) and in model 4 with duration of surgery (AUC 0.85). In each case, a score was generated that allowed an indication for the EPOCT to be determined. It was shown that neurological deficits had a strong correlation with the result of the EPOCT. In view of a possible indication for an emergency return to the operating room, it was shown that the neurological examination was a better predictor than the EPOCT. In no single case was an emergency reoperation indicated by abnormal imaging alone. In summary, it can be concluded that routine EPOCT is not superior to conventional neurological examination for the indication of emergency reoperation after neurosurgical tumor resection. Risk factors were identified that were proven to be significantly associated with abnormal control imaging or a change in patient management. In addition, scores were created that enable an assessment of the outcome of the EPOCT in causal relation with two variables. As things stand, it is not yet possible to create an exact score that can serve as a guide across all cases. Until this is achieved, ideally through a larger, prospective study, the EPOCT must be removed from its position as a routine control and used merely as a supportive tool.10000-01-01Diese Dateien sind bis zum Abschluß der Staatsexamensprüfung gesperrt.These files are not accessible until the state exam has been completed