Multiple myeloma (MM) is consistently preceded by precursor conditions recognized clinically as monoclonal gammopathy of undetermined significance (MGUS) or smoldering myeloma (SMM). We interrogate the whole genome sequence (WGS) profile of 18 MGUS and compare them with those from 14 SMMs and 80 MMs. We show that cases with a non-progressing, clinically stable myeloma precursor condition (n = 15) are characterized by later initiation in the patient’s life and by the absence of myeloma defining genomic events including: chromothripsis, templated insertions, mutations in driver genes, aneuploidy, and canonical APOBEC mutational activity. This data provides evidence that WGS can be used to recognize two biologically and clinically distinct myeloma precursor entities that are either progressive or stable.

The flatworm species Schmidtea mediterranea and Macrostomum lignano have become new and innovative model organisms in stem cell, regeneration and tissue homeostasis research. Because of their unique stem cell system, (lab) technical advantages and their phylogenetic position within the Metazoa, they are also ideal candidate model organisms for toxicity assays. As stress and biomarker screenings are often performed at the transcriptional level, the aim of this study was to establish a set of reference genes for qPCR experiments for these two model organisms in different stress situations. We examined the transcriptional stability of nine potential reference genes (actb, tubb, ck2, cox4, cys, rpl13, gapdh, gm2ap, plscr1) to assess those that are most stable during altered stress conditions (exposure to carcinogenic metals and salinity stress). The gene expression stability was evaluated by means of geNorm and NormFinder algorithms. Sets of best reference genes in these analyses varied between different stress situations, although gm2ap and actb were stably transcribed during all tested combinations. In order to demonstrate the impact of bad normalisation, the stress-specific gene hsp90 was normalised to different sets of reference genes. In contrast to the normalisation according to GeNorm and NormFinder, normalisation of hsp90 in Macrostomum lignano during cadmium stress did not show a significant difference when normalised to only gapdh. On the other hand an increase of variability was noticed when normalised to all nine tested reference genes together. Testing appropriate reference genes is therefore strongly advisable in every new experimental condition.

Genomic screening studies recently revealed that mutations in ribosomal protein (RP) genes represent a novel class of defects in cancer. In T-cell acute lymphoblastic leukemia (T-ALL), 20% of children harbor acquired mutations and deletions in RPL10 (uL16 in the new nomenclature 1 ), RPL5 (uL18) and RPL22 (eL22), 3 proteins of the large 60S ribosomal subunit.2,3 Strikingly, 7.9% of pediatric T-ALL patients carried the same RPL10 R98S missense mutation.2 Somatic mutations in RPs are not confined to T-ALL. RPL5 is mutated in 11-34% of glioblastoma, melanoma and breast cancer samples, and 10-20% of chronic lymphocytic leukemia samples have RPS15 mutations. 4,5,6 The plasma cell malignancy multiple myeloma (MM) is an attractive candidate for harboring RP mutations: initial genome sequencing revealed that half of the patients carry mutations in genes that may be functionally linked to protein translation, and we recently described that RPL5 is in a 58 kb minimal deleted region on 1p22 that is deleted in ≥20% of MM cases.

The use of targeted Next Generation Sequencing (NGS) for the diagnostic screening of somatic variants in solid tumor samples has proven its high clinical value. Because of the large number of ongoing clinical trials for a multitude of variants in a growing number of genes, as well as the detection of proven and emerging pan-cancer biomarkers including microsatellite instability (MSI) and tumor mutation burden (TMB), the currently employed diagnostic gene panels will become vastly insufficient in the near future. Here, we describe the validation and implementation of the hybrid capture-based comprehensive TruSight Oncology (TSO500) assay that is able to detect single-nucleotide variants (SNVs) and subtle deletions and insertions (indels) in 523 tumor-associated genes, copy-number variants (CNVs) of 69 genes, fusions with 55 cancer driver genes, and MSI and TMB. Extensive validation of the TSO500 assay was performed on DNA or RNA from 170 clinical samples with neoplastic content down to 10%, using multiple tissue and specimen types. Starting with 80 ng DNA and 40 ng RNA extracted from formalin-fixed and paraffine-embedded (FFPE) samples revealed a precision and accuracy >99% for all variant types. The analytical sensitivity and specificity were at least 99% for SNVs, indels, CNVs, MSI, and gene rearrangements. For TMB, only values around the threshold could yield a deviating outcome. The limit-of-detection for SNVs and indels was well below the set threshold of 5% variant allele frequency (VAF). This validated comprehensive genomic profiling assay was then used to screen 624 diagnostic samples, and its success rate for adoption in a clinical diagnostic setting of broad solid tumor screening was assessed on this cohort.