Core facility for molecular cytogenetics, genetic diagnostics and profiling

在上一个资助期间，我们在临床研究单位（CRU）内建立了集中样本收集。来自<s:1>茨堡大学和乌尔姆大学的患者样本已被表征为细胞遗传学和分子遗传学的变化。CD138纯化的浆细胞被用于肿瘤样品载玻片和肿瘤细胞颗粒的存档，以供进一步分析。另外，将相应的种系材料存档作为对照。使用SNP芯片分析（Affymetrix Microarray Human Genome SNP 6.0）对MM样品进行高分辨率全基因组遗传改变筛查。在下一个资助期内，我们将扩大临床样本的收集。从德国多发性骨髓瘤研究小组（DSMM）开展的临床试验中纳入特征明确的患者，将加强肿瘤库，并将成为CRU内转化研究的宝贵资源。最近发表的通过大规模平行测序对38个MM肿瘤基因组突变景观的表征突出了多发性骨髓瘤发病机制的新方面，包括检测新的突变候选基因。因此，我们将特别关注通过使用新的下一代测序（NGS）技术对初级MM样品进行精细的分子表征。从参与DSMM试验的非常有特征的患者的许多MM样本中，我们将对一组全面的靶基因进行突变筛选：NRAS、KRAS、BRAF、TP53、DIS3、FAM46C、KDM6A[也称为UTX]、TRAF3、CYLD、CDKN2A/CDKN2B和CDKN2C。这项研究将伴随着转录组测序（RNA-seq）方法，以便对DNA变异和基因表达变化进行综合分析。此外，RNA-seq数据提供了检测mm相关融合基因以及异常/差异剪接模式的可能性，这些剪接模式最近也被证明与发病相关。同样，在过去的十年中，小的非编码rna，特别是microrna （miRNA）被确定为正常和病理造血的关键调节因子。然而，mirna在浆细胞疾病发展中的作用以及导致MM发展的致癌途径尚不完全清楚。因此，我们还将尝试确定CRU项目研究的各种MM亚组中mirna的差异表达模式。

During the last funding period we have established a centralized sample collection within the Clinical Research Unit (CRU). Patient samples from the University of Würzburg as well as from the University of Ulm have been characterized for cytogenetic and molecular genetic changes. The CD138 purified plasma cells are processed for archiving of tumor sample slides and of tumor cell pellets for further analysis. Additionally, corresponding germline material is archived as reference control. High-resolution genome-wide screening for genetic alterations of MM samples was performed using SNP chip analysis (Affymetrix Microarray Human Genome SNP 6.0). During the next funding period we will expand the collection of clinical samples. The incorporation of well characterized patients from clinical trials run by the German Study Group on Multiple Myeloma (DSMM) will strengthen the tumor bank and will serve as an invaluable resource for translational research within the CRU. The recently published characterization of the mutational landscape of 38 MM tumor genomes through massively parallel sequencing highlighted new aspects of the pathogenesis of multiple myeloma including the detection of novel mutated candidate genes.Consequently, we will put a special focus on a refined molecular characterization of the primary MM samples by the use of novel next-generation sequencing (NGS) technologies. From a number of MM samples from very well characterized patients participating in DSMM trials we will perform a mutational screen in a comprehensive set of target genes: NRAS, KRAS, BRAF, TP53, DIS3, FAM46C, KDM6A [also known as UTX], TRAF3, CYLD, CDKN2A/CDKN2B and CDKN2C. This research will be accompanied by a transcriptome sequencing (RNA-seq) approach in order to allow integrative analyses of DNA variation and gene expression changes. Furthermore, RNA-seq data offers the possibility to detect MMassociated fusion genes as well as aberrant/differential splicing patterns which recently have been shown to be also of pathogenetic relevance. Similarly, over the last decade small noncoding RNAs, particularly microRNAs (miRNA) were identified as key regulators of normal and pathological hematopoiesis. However, the role of miRNAs in the development of plasma cell disorders and the oncogenic pathways that lead to the development of MM is not fully understood. Therefore, we will also try to identify differential expression patterns of miRNAs in various MM subgroups investigated by the CRU projects.