Genetische Subgruppen und individuelle Krankheitsursache der Akuten Myeloischen Leukämie (AML)

Neuartige Sequenzierverfahren ermöglichen die systematische Identifikation von somatischen Mutationen in Tumorproben. Mittels Exomsequenzierung haben wir Subgruppen der Akuten Myeloischen Leukämie (AML) untersucht und spezifische Assoziationen von Mutationen entdeckt. Beispielsweise fanden wir Läsionen im CEBPA-GATA2 Transkriptionsfaktor-Komplex, welche die Aktivierung von Zielgenen stören. Außerdem identifizierten wir in der AML M4eo Subgruppe die N676K Mutation im FLT3-Rezeptor, deren wachstumssteigernder Effekt mit Tyrosinkinase-Inhibitoren gehemmt werden kann. AML Patienten mit Trisomie 13 zeigten einen sehr ungünstigen klinischen Verlauf sowie ein homogenes Mutationsprofil mit häufigen Mutationen in Spliceosomgenen. Bei der AML mit t(8;21) Translokation fanden wir häufige genetische Läsionen in ZBTB7A, die zu einer Deregulation des Tumorstoffwechsels führen. Weiterhin detektierten wir in den Sequenzdaten somatische Copy Number Alterations (CNAs), welche die zytogenetischen Befunde in der Routinediagnostik ergänzen können. Die Veränderung der Mutationsprofile im Krankheitsverlauf war mit dem Rezidiv-freien Überleben korreliert und wies auf Mechanismen der Therapieresistenz hin. Das Verständnis der individuellen Krankheitsursache der AML liefert neue pharmakologische Angriffspunkte und bereitet den Weg für maßgeschneiderte Behandlungen

Charakterisierung von Chromatinfaktoren mit Hilfe von Antikörpern: Von der Fruchtfliege zum humanen Prostatakarzinom und zurück

Die Organisation der DNA in Nukleosomen hat einen großen Einfluss auf die Regulation von grundliegenden Prozessen wie Transkription, Replikation oder Reparatur der DNA im Zellkern. Um die hinderliche Natur des Chromatins bei diesen fundamentalen Prozessen zu überwinden, existieren mehrere verschiedene Chromatin modifizierende Proteinkomplexe im Zellkern. Chromatin Remodelling Komplexe nützen die Energie der ATP-Hydrolyse um die Position der Nukleosomen so zu verändern, dass verschiedene Abschnitte der DNA für die Interaktion mit regulierenden Faktoren zugänglich werden. Ein Klasse solcher Remodelling Faktoren beinhalten die ATPase ISWI als katalytische Untereinheit. Das Protein wurde zuerst in Drosophila entdeckt und die drei verschiedenen ISWI enthaltenden Komplexe, nämlich NURF, ACF und CHRAC, wurden ausführlich in diesem Modellorganismus untersucht. Homolog zur Fruchtfliege existieren sehr ähnliche Protein Komplexe beim Menschen. Wir haben das humane ISWI mit den Isoformen Snf2h und Snf2L im Prostatakarzinom untersucht. In einem Tissue Microarray wurden Gewebeproben mit Hilfe von polyklonalen Antikörpern gegen ISWI gefärbt. Es folgte ein quantitativer Vergleich der Färbungsintensitäten im Karzinomgewebe sowie in gutartigem Gewebe der Prostata durch Anwendung von digitaler Bildanalyse. Das Ergebnis war eine signifikant stärkere Färbung im neoplastischen Gewebe. Eine Anreicherung von ISWI in Krebszellen ist besonders interessant im Kontext der bekannten Funktionen des Proteins für DNA-Replikation, Zellproliferation und Regulation der Chromatinstruktur. In einem zweiten Projekt sind wir zum Modell der Fruchtfliege zurückgekehrt und entwickelten monoklonale Antikörper gegen Toutatis, das zu einer Proteinfamilie gehört, die auch einige bekannte Interaktionspartner von ISWI umfasst. Die Proteine dieser Familie haben vermutlich eine regulatorische Funktion in den Remodelling Komplexen, denn am Beispiel von Acf1 wurde gezeigt, dass sie die nukleosomale Bindung sowie die Effizienz und Richtung der Mobilisierung von Nukleosomen modifizieren. Unsere Antikörper wurden etabliert, um Toutatis enthaltende Komplexe durch Western Blot Analyse von gereinigten Drosophila-Extrakten und Immunfluoreszenz zu charakterisieren. Mit diesen Methoden fanden wir eine Koelution von Toutatis mit der ATPase Brahma und dem Strukturprotein Spectrin alpha sowie eine Lokalisation in der Lamina des Zellkerns. Ein mögliches Zusammenspiel dieser Proteine in einem neuen Chromatin Remodelling Komplex mit einer Beteiligung an der DNA-Reparatur wird diskutiert

The Success of Others: Copy Trading and Risk

Social trading platforms and day trading apps are increasingly popular. They allow copy trading which means that trades of another trader are automatically copied. Traders can provide credentials, which has been shown to make them seem trustworthy. These features have been shown to increase the tendency of inexperienced users to take risks. We present the design and preliminary results of an experimental study that investigates whether the negative externalities of social trading can be alleviated if the corresponding trades have to be executed actively. In the experiment, the users observe the trading behavior and success of anonymous other users and are then asked to take investment decisions without being able to automatically trade as the observed investors. Pretest results indicate that having to execute the investment decisions themselves might reduce the tendency to take risky decisions. This is a promising avenue for the regulation of day trading apps

GATA2 mutations in myeloid malignancies: Two zinc fingers in many pies

The GATA family of transcription factors are zinc finger (ZF) DNA‐binding proteins that regulate transcription during development and cell differentiation. GATA2 plays an essential role in the regulation of hematopoiesis. As a result, mutations in this gene or alterations in its expression level or function have been linked to a variety of human hematologic disorders. In this review, we summarize the findings and developments over the recent years regarding the clinical correlations and functional properties of distinct GATA2 mutations in hematopoietic malignancies, with particular focus on the mutational hotspots in the ZF domains

Diagnostic applications of next generation sequencing: working towards quality standards

Over the past 6 years, next generation sequencing (NGS) has been established as a valuable high-throughput method for research in molecular genetics and has successfully been employed in the identification of rare and common genetic variations. All major NGS technology companies providing commercially available instruments (Roche 454, Illumina, Life Technologies) have recently marketed bench top sequencing instruments with lower throughput and shorter run times, thereby broadening the applications of NGS and opening the technology to the potential use for clinical diagnostics. Although the high expectations regarding the discovery of new diagnostic targets and an overall reduction of cost have been achieved, technological challenges in instrument handling, robustness of the chemistry and data analysis need to be overcome. To facilitate the implementation of NGS as a routine method in molecular diagnostics, consistent quality standards need to be developed. Here the authors give an overview of the current standards in protocols and workflows and discuss possible approaches to define quality criteria for NGS in molecular genetic diagnostics

Transcutaneous aortic valve replacement with the Edwards SAPIEN XT and Medtronic CoreValve prosthesis under fluoroscopic guidance and local anaesthesia only

Objective: To assess the feasibility of transcatheter aortic valve implantation (TAVI; Medtronic CoreValve and Edwards SAPIEN XT) under local anaesthesia with only mild analgesic medication and fluoroscopic guidance. Methods: 461 patients underwent TAVI under local anaesthesia with lidocaine. The procedure was performed successfully in 459 of the cases. All patients were also treated with piritramide, metoclopramide hydrochloride and 62 mg dimenhydrinate. Monitoring consisted of a six-electrode, virtual 12-lead ECG, pulse oximetry, and invasive arterial pressure measurement. There was no continuous surveillance by an anaesthesiologist. Results: There was no need for conversion to general anaesthesia except in four patients who required cardiopulmonary resuscitation. Conscious sedation with intravenous administration of midazolam for agitation or inotropic medication for prolonged hypotension was necessary in only seven of the 461 patients. The combined safety end point according to the Valve Academic Research Consortium consensus document was reached in 12.6%. Conclusions: Our results show that TAVI performed under local anaesthesia with only mild analgesic medication and under fluoroscopic guidance is feasible, with good outcome comparable to published data

Applications and data analysis of next-generation sequencing

Over the past 6 years, next-generation sequencing (NGS) has been established as a valuable high-throughput method for research in molecular genetics and has successfully been employed in the identification of rare and common genetic variations. Although the high expectations regarding the discovery of new diagnostic targets and an overall reduction of cost have been achieved, technological challenges in instrument handling, robustness of the chemistry, and data analysis need to be overcome. Each workflow and sequencing platform have their particular problems and caveats, which need to be addressed. Regarding NGS, there is a variety of different enrichment methods, sequencing devices, or technologies as well as a multitude of analyzing software products available. In this manuscript, the authors focus on challenges in data analysis when employing different target enrichment methods and the best applications for each of the

NGS-guided precision oncology in metastatic breast and gynecological cancer: first experiences at the CCC Munich LMU

PURPOSE Comprehensive genomic profiling identifying actionable molecular alterations aims to enable personalized treatment for cancer patients. The purpose of this analysis was to retrospectively assess the impact of personalized recommendations made by a multidisciplinary tumor board (MTB) on the outcome of patients with breast or gynecological cancers, who had progressed under standard treatment. Here, first experiences of our Comprehensive Cancer Center Molecular Tumor Board are reported. METHODS All patients were part of a prospective local registry. 95 patients diagnosed with metastatic breast cancer or gynecological malignancies underwent extended molecular profiling. From May 2017 through March 2019, the MTB reviewed all clinical cases considering tumor profile and evaluated molecular alterations regarding further diagnostic and therapeutic recommendations. RESULTS 95 patients with metastatic breast or gynecological cancers were discussed in the MTB (68% breast cancer, 20% ovarian cancer, 5% cervical cancer, 3% endometrial cancer and 4% others). Genes with highest mutation rate were PIK3CA and ERBB2. Overall, 34 patients (36%) received a biomarker-based targeted therapy recommendation. Therapeutic recommendations were implemented in nine cases; four patients experienced clinical benefit with a partial response or disease stabilization lasting over 4~months. CONCLUSION In the setting of a multidisciplinary molecular tumor board, a small but clinically meaningful group of breast and gynecological cancer patients benefits from comprehensive genomic profiling. Broad and successful implementation of precision medicine is complicated by patient referral at late stage disease and limited access to targeted agents and early clinical trials. TRIAL REGISTRATION NUMBER 284-10 (03.05.2018)

ZBTB7A prevents RUNX1-RUNX1T1-dependent clonal expansion of human hematopoietic stem and progenitor cells

ZBTB7A is frequently mutated in acute myeloid leukemia (AML) with t(8;21) translocation. However, the oncogenic collaboration between mutated ZBTB7A and the RUNX1–RUNX1T1 fusion gene in AML t(8;21) remains unclear. Here, we investigate the role of ZBTB7A and its mutations in the context of normal and malignant hematopoiesis. We demonstrate that clinically relevant ZBTB7A mutations in AML t(8;21) lead to loss of function and result in perturbed myeloid differentiation with block of the granulocytic lineage in favor of monocytic commitment. In addition, loss of ZBTB7A increases glycolysis and hence sensitizes leukemic blasts to metabolic inhibition with 2-deoxy-d-glucose. We observed that ectopic expression of wild-type ZBTB7A prevents RUNX1-RUNX1T1-mediated clonal expansion of human CD34+ cells, whereas the outgrowth of progenitors is enabled by ZBTB7A mutation. Finally, ZBTB7A expression in t(8;21) cells lead to a cell cycle arrest that could be mimicked by inhibition of glycolysis. Our findings suggest that loss of ZBTB7A may facilitate the onset of AML t(8;21), and that RUNX1-RUNX1T1-rearranged leukemia might be treated with glycolytic inhibitors