Targetclone: A multi-sample approach for reconstructing subclonal evolution of tumors

Most tumors are composed of a heterogeneous population of subclones. A more detailed insight into the subclonal evolution of these tumors can be helpful to study progression and treatment response. Problematically, tumor samples are typically very heterogeneous, making deconvolving individual tumor subclones a major challenge. To overcome this limitation, reducing heterogeneity, such as by means of microdissections, coupled with targeted sequencing, is a viable approach. However, computational methods that enable reconstruction of the evolutionary relationships require unbiased read depth measurements, which are commonly challenging to obtain in this setting. We introduce TargetClone, a novel method to reconstruct the subclonal evolution tree of tumors from single-nucleotide polymorphism allele frequency and somatic single-nucleotide variant measurements. Furthermore, our method infers copy numbers, alleles and the fraction of the tumor component in each sample. TargetClone was specifically designed for targeted sequencing data obtained from microdissected samples. We demonstrate that our method obtains low error rates on simulated data. Additionally, we show that our method is able to reconstruct expected trees in a testicular germ cell cancer and ovarian cancer dataset. The TargetClone package including tree visualization is written in Python and is publicly available at https://github.com/UMCUGenetics/targetclone

Origin and clinical relevance of chromosomal aberrations other than the common trisomies detected by genome-wide NIPS: Results of the TRIDENT study

PurposeNoninvasive prenatal screening (NIPS) using cell-free DNA in maternal blood is highly sensitive for detecting fetal trisomies 21, 18, and 13. Using a genome-wide approach, other chromosome anomalies can also be detected. We report on the origin

Mosaic maternal 10qter deletions are associated with FRA10B expansions and may cause false-positive noninvasive prenatal screening results

Purpose: Using genome-wide noninvasive prenatal screening (NIPS), we detected a 20-megabase specific deletion starting at 10q25 in eight pregnancies. The deletion could not be confirmed by invasive testing. Since all 10(q25→qter) deletions started close to the FRA10B fragile site in 10q25, we investigated whether the pregnant women were indeed carriers of FRA10B. Methods: We performed NIPS analysis for all autosomes using single-read sequencing. Analysis was done with the WISECONDOR algorithm. Culture of blood lymphocytes with bromodeoxyuridine was used to detect FRA10B expansions. Fluorescence in situ hybridization and array analysis were used to find maternal and/or fetal deletions. Results: We confirmed the presence of a FRA10B expansion in all four tested mothers. Fluorescence in situ hybridization and array analysis confirmed the presence of a maternal mosaic deletion of 10(q25→qter). Conclusion: The recurring 10(q25→qter) deletion detected with NIPS is a false-positive result caused by a maternal low-level mosaic deletion associated with FRA10B expansions. This has important consequences for clinical follow-up, as invasive procedures are unnecessary. Expanded maternal FRA10B repeats should be added to the growing group of variants in the maternal genome that may cause false-positive NIPS results

WISExome: A within-sample comparison approach to detect copy number variations in whole exome sequencing data

In clinical genetics, detection of single nucleotide polymorphisms (SNVs) as well as copy number variations (CNVs) is essential for patient genotyping. Obtaining both CNV and SNV information from WES data would significantly simplify clinical workflow. Unfortunately, the sequence reads obtained with WES vary between samples, complicating accurate CNV detection with WES. To avoid being dependent on other samples, we developed a within-sample comparison approach (WISExome). For every (WES) target region on the genome, we identified a set of reference target regions elsewhere on the genome with similar read frequency behavior. For a new sample, aberrations are detected by comparing the read frequency of a target region with the distribution of read frequencies in the reference set. WISExome correctly identifies known pathogenic CNVs (range 4 Kb–5.2 Mb). Moreover, WISExome prioritizes pathogenic CNVs by sorting them on quality and annotations of overlapping genes in OMIM. When comparing WISExome to four existing CNV detection tools, we found that CoNIFER detects much fewer CNVs and XHMM breaks calls made by other tools into smaller calls (fragmentation). CODEX and CLAMMS seem to perform more similar to WISExome. CODEX finds all known pathogenic CNVs, but detects much more calls than all other methods. CLAMMS and WISExome agree the most. CLAMMS does, however, miss one of the known CNVs and shows slightly more fragmentation. Taken together, WISExome is a promising tool for genome diagnostics laboratories as the workflow can be solely based on WES data.Pattern Recognition and Bioinformatic

Enhancer hubs and loop collisions identified from single-allele topologies

Chromatin folding contributes to the regulation of genomic processes such as gene activity. Existing conformation capture methods characterize genome topology through analysis of pairwise chromatin contacts in populations of cells but cannot discern whether individual interactions occur simultaneously or competitively. Here we present multi-contact 4C (MC-4C), which applies Nanopore sequencing to study multi-way DNA conformations of individual alleles. MC-4C distinguishes cooperative from random and competing interactions and identifies previously missed structures in subpopulations of cells. We show that individual elements of the β-globin superenhancer can aggregate into an enhancer hub that can simultaneously accommodate two genes. Neighboring chromatin domain loops can form rosette-like structures through collision of their CTCF-bound anchors, as seen most prominently in cells lacking the cohesin-unloading factor WAPL. Here, massive collision of CTCF-anchored chromatin loops is believed to reflect ‘cohesin traffic jams’. Single-allele topology studies thus help us understand the mechanisms underlying genome folding and functioning

Determination of alpha-s for b quarks at the Z0 resonance

The strong coupling constant for b quarks has been determined, and its flavour independence, as predicted by QCD, investigated. The analysis involved events with lepton candidates selected from approximately 356 000 hadronic decays of the Z0, collected by the DELPHI detector at LEP in 1990 and 1991. A method based on a direct comparison of the three-jet fraction in a b enriched sample, selected by requiring leptons with large momenta, to that of the entire hadronic sample, illustrated the significant effect of the b quark mass on the multi-jet cross section, and verified the flavour independence of the strong coupling constant to an accuracy of ±6%. A second procedure based on a fit to the momentum and transverse momentum spectra of the lepton candidates in both two-jet and three (or more)-jet event samples simultaneously determined the b content in each, and, using second order QCD calculations, gave an absolute measurement of αs for b quarks of 0.118±0.004 (stat.)±0.003 (syst.)±0.008 (scale). A comparison with αs for all quark flavours, as measured from the three-jet fraction in all hadronic events, further allowed the coupling strength for b quarks to be expressed in terms relative to that for udsc quarks, thereby cancelling certain common systematic uncertainties, and yielded View the MathML source