Innovative Therapiekonzepte in mesenchymalen Neoplasien

Sarkome sind eine Gruppe heterogener Tumore, deren Therapieoptionen überwiegend in konventioneller Chemo- oder Strahlentherapie bestehen. Ansätze einer personalisierten Medizin sind nur bei wenigen Subentitäten etabliert. Jedoch besteht ein hoher Bedarf an alternativen Therapieoptionen für klinisch aggressive Sarkomen sowie für Tumore, die unter Therapie progredient sind. Ansätze dafür sind miRNAs, HSP90 Inhibitoren, Inhibitoren epigenetischer Schlüsselfaktoren wie Histon-Deacetylasen, Histon- Acetyltransferasen und DNA Methyltransferasen, aber auch Inhibitoren der Zellzykluskontrolle. Die Expression von HR23b ist ein prädiktiver Biomarker für die Sensitivität gegenüber HDACi in kutanen T Zell Lymphomen und hepatozellulären Karzinomen. Im Rahmen dieser Arbeit wurden HDACi und der Einsatz von miRNAs als alternative Therapieoptionen in mesenchymalen Neoplasien untersucht. Als Modelsystem diente ein Kollektiv aus 17 verschiedenen Sarkom- und GIST-Zelllinien, das alle wichtigen Sarkomentitäten abdeckte, die bei Erwachsenen auftreten können. Westernblot Analysen zeigten, dass die ausgewählten Sarkom- und GIST-Zelllinien ein breites Spektrum an HR23b Expression aufwiesen. Die HDACi Behandlung mit vier verschiedenen HDACi zeigte in Sarkomen antiproliferative und proapoptotische Effekte. Eine signifikante Korrelation zwischen HR23b Expression und Sensitivität gegenüber Vorinostat konnte gezeigt werden. Die HR23b Expression wurde konzentrationsabhängig unter HDACi reduziert. In einem Kollektiv aus über 500 klinischen Sarkom- und GIST-Proben wiesen 12,5% der Sarkome und 23,2% der GISTs eine starke HR23b Expression auf. Vor allem aggressive Entitäten wie maligne periphere Nervenscheidewandtumore, Leiomyosarkome und Angiosarkome zeigten eine hohe HR23b Positivität, die sie zu potentiellen neuen Kandidaten für weitere Studien mit HDACi basierter Therapie machen. Aber auch miRNAs stellen eine neue potentielle Therapieoption in der Behandlung von Tumoren dar. In der vorliegenden Arbeit wurden drei verschieden GIST-Zelllinien verwendet, um den miR-221 und miR-222 vermittelten Effekt in Bezug auf die Proliferation, Apoptose und Signaltransduktion zu analysieren. MTT und ApoTox-GloTM Triplex Assays zeigten, dass die miRNAs die zelluläre Proliferation in allen drei GIST Zelllinien reduzierten und dass diese Reduktion mit einer signifikanten Apoptoseinduktion korrelierte. Die Effekte der miRNAs waren weder additiv noch kompetitiv. Westernblot Analysen zur Signaltransduktion zeigten, dass diese antiproliferativen und proapoptotischen Effekte über eine Signalkaskade von KIT, AKT und BCL2, aber nicht über MTOR und BCL2L11 vermittelt wurde

The impact of sequencing on diagnosis and treatment of malignant melanoma

Melanoma is one of the clinically most important cancer types considering its high mortality rate and that it is commonly diagnosed in relatively young people. With the advent of targeted therapies and, more recently, immune checkpoint inhibitors, more treatment options are available resulting in higher patient survival rates. However, the successful application of these targeted therapies critically depends on the reliable detection of molecular aberrations. Today, massively parallel sequencing techniques enable us to analyze large sets of genes in a relatively short time. It has allowed increased knowledge of acquired somatic mutations in melanoma and has helped to identify new targets for personalized therapy, and potentially may help to predict response to immune therapies. Described here are the development of sequencing techniques, how their improvement has changed diagnosis, prognosis and management of malignant melanoma and the future perspectives of melanoma diagnostics in the routine clinical setting

MET gene copy number alterations and expression of MET and hepatocyte growth factor are potential biomarkers in angiosarcomas and undifferentiated pleomorphic sarcomas.

Soft tissue sarcomas are a heterogeneous group of tumors with many different subtypes. In 2014 an estimated 12,020 newly diagnosed cases and 4,740 soft tissue sarcoma related deaths can be expected in the United States. Many soft tissue sarcomas are associated with poor prognosis and therapeutic options are often limited. The evolution of precision medicine has not yet fully reached the clinical treatment of sarcomas since therapeutically tractable genetic changes have not been comprehensively studied so far. We analyzed a total of 484 adult-type malignant mesenchymal tumors by MET fluorescence in situ hybridization and MET and hepatocyte growth factor immunohistochemistry. Eleven different entities were included, among them the most common and clinically relevant subtypes and tumors with specific translocations or complex genetic changes. MET protein expression was observed in 2.6% of the cases, all of which were either undifferentiated pleomorphic sarcomas or angiosarcomas, showing positivity rates of 14% and 17%, respectively. 6% of the tumors showed hepatocyte growth factor overexpression, mainly seen in undifferentiated pleomorphic sarcomas and angiosarcomas, but also in clear cell sarcomas, malignant peripheral nerve sheath tumors, leiomyosarcomas and gastrointestinal stromal tumors. MET and hepatocyte growth factor overexpression were significantly correlated and may suggest an autocrine activation in these tumors. MET FISH amplification and copy number gain were present in 4% of the tumors (15/413). Two samples, both undifferentiated pleomorphic sarcomas, fulfilled the criteria for high level amplification of MET, one undifferentiated pleomorphic sarcoma reached an intermediate level copy number gain, and 12 samples of different subtypes were categorized as low level copy number gains for MET. Our findings indicate that angiosarcomas and undifferentiated pleomorphic sarcomas rather than other frequent adult-type sarcomas should be enrolled in screening programs for clinical trials with MET inhibitors. The screening methods should include both in situ hybridization and immunohistochemistry

Genetic instability and recurrent MYC amplification in ALK-translocated NSCLC: a central role of TP53 mutations

The anaplastic lymphoma kinase (ALK) rearrangement defines a distinct molecular subtype of non-small cell lung cancer (NSCLC). Despite the excellent initial efficacy of ALK inhibitors in patients with ALK+ lung cancer, resistance occurs almost inevitably. To date, there is no reliable biomarker allowing the identification of patients at higher risk of relapse. Here, we analysed a subset of 53 ALK+ tumors with and without TP53 mutation and ALK+ NSCLC cell lines by NanoString nCounter technology. We found that the co-occurrence of early TP53 mutations in ALK+ NSCLC can lead to chromosomal instability: 24% of TP53-mutated patients showed amplifications of known cancer genes such as MYC (14%), CCND1 (10%), TERT (5%), BIRC2 (5%), ORAOV1 (5%), and YAP1 (5%). MYC-overexpressing ALK+ TP53-mutated cells had a proliferative advantage compared to wild-type cells. ChIP-Seq data revealed MYC-binding sites within the promoter region of EML4, and MYC overexpression in ALK+ TP53-mutated cells resulted in an upregulation of EML4-ALK, indicating a potential MYC-dependent resistance mechanism in patients with increased MYC copy number. Our study reveals that ALK+ NSCLC represents a more heterogeneous subgroup of tumors than initially thought, and that TP53 mutations in that particular cancer type define a subset of tumors that harbour chromosomal instability, leading to the co-occurrence of pathogenic aberrations. (c) 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland

MET and HGF expressions.

<p>Representative sarcomas showing range and pattern of immunohistochemical staining for MET (A-H) and HGF (I-P). (A, B, I, J) staining intensity score = 0, (C, D, K, L) score = 1+, (E, F, G, M, N, O,) score = 2+, (H, P) score = 3+. (A, C, E, N) undifferentiated pleomorphic sarcomas, (B, D, F, G, K, M) angiosarcomas, (H) control cell line, (I, L, O) leiomyosarcomas, (J) dedifferentiated liposarcoma, (P) control tissue breast cancer.</p

FISH results of <i>MET</i> positive cases.

<p>ASA, angiosarcoma; CCS, clear cell sarcoma; CEN, centromere; DDLS, dedifferentiated liposarcoma; GIST, gastrointestinal stromal tumor; ID, identification; LMS, leiomyosarcoma; MFH, undifferentiated pleomorphic sarcoma; MLS, myxoid liposarcomas; PLS, pleomorphic liposarcoma</p><p>FISH results of <i>MET</i> positive cases.</p

Fluorescence <i>in situ</i> hybridization for <i>MET</i>.

<p>Representative sarcomas showing different categories for <i>MET</i> gene copy number variations (see text for explanation). <i>MET</i> gene is labeled in green, centromere 7 in orange. (A) <i>MET</i> negative clear cell sarcoma, (B) <i>MET</i> negative angiosarcoma, (C) low level copy number gain in a clear cell sarcoma, (D) intermediate level copy number gain in an undifferentiated pleomorphic sarcoma, (E, F) high level amplification in undifferentiated pleomorphic sarcomas.</p

ClinicalandPathological Characteristics of KEAP1- and NFE2L2-Mutated Non-Small Cell Lung Carcinoma (NSCLC)

Purpose: KEAP1 and NFE2L2 mutations are associated with impaired prognosis in a variety of cancers and with squamous cell carcinoma formation in non-small cell lung cancer (NSCLC). However, little is known about frequency, histology dependence, molecular and clinical presentation as well as response to systemic treatment in NSCLC. Experimental Design: Tumor tissue of 1,391 patients with NSCLC was analyzed using next-generation sequencing (NGS). Clinical and pathologic characteristics, survival, and treatment outcome of patients with KEAP1 or NFE2L2 mutations were assessed. Results: KEAP1 mutations occurred with a frequency of 11.3% (n = 157) and NFE2L2 mutations with a frequency of 3.5% (n = 49) in NSCLC patients. In the vast majority of patients, both mutations did not occur simultaneously. KEAP1 mutations were found mainly in adenocarcinoma (AD; 72%), while NFE2L2 mutations were more common in squamous cell carcinoma (LSCC; 59%). KEAP1 mutations were spread over the whole protein, whereas NFE2L2 mutations were clustered in specific hotspot regions. In over 80% of the patients both mutations co-occurred with other cancer-related mutations, among them also targetable aberrations like activating EGFR mutations or MET amplification. Both patient groups showed different patterns of metastases, stage distribution and performance state. No patient with KEAP1 mutation had a response on systemic treatment in first-, second-, or third-line setting. Of NFE2L2-mutated patients, none responded to second-or third-line therapy. Conclusions: KEAP1- and NFE2L2-mutated NSCLC patients represent a highly heterogeneous patient cohort. Both are associated with different histologies and usually are found together with other cancer-related, partly targetable, genetic aberrations. In addition, both markers seem to be predictive for chemotherapy resistance. (C) 2018 AACR