Timo Miettinen

Comparison of solution-based exome capture methods for next generation sequencing

BackgroundTechniques enabling targeted re-sequencing of the protein coding sequences of the human genome on next generation sequencing instruments are of great interest. We conducted a systematic comparison of the solution-based exome capture kits provided by Agilent and Roche NimbleGen. A control DNA sample was captured with all four capture methods and prepared for Illumina GAII sequencing. Sequence data from additional samples prepared with the same protocols were also used in the comparison.ResultsWe developed a bioinformatics pipeline for quality control, short read alignment, variant identification and annotation of the sequence data. In our analysis, a larger percentage of the high quality reads from the NimbleGen captures than from the Agilent captures aligned to the capture target regions. High GC content of the target sequence was associated with poor capture success in all exome enrichment methods. Comparison of mean allele balances for heterozygous variants indicated a tendency to have more reference bases than variant bases in the heterozygous variant positions within the target regions in all methods. There was virtually no difference in the genotype concordance compared to genotypes derived from SNP arrays. A minimum of 11× coverage was required to make a heterozygote genotype call with 99% accuracy when compared to common SNPs on genome-wide association arrays.ConclusionsLibraries captured with NimbleGen kits aligned more accurately to the target regions. The updated NimbleGen kit most efficiently covered the exome with a minimum coverage of 20×, yet none of the kits captured all the Consensus Coding Sequence annotated exons.

Robust denoising of electrophoresis and mass spectrometry signals with minimum description length principle

The need for high‐throughput assays in molecular biology places increasing requirements on the applied signal processing and modelling methods. In order to be able to extract useful information from the measurements, the removal of undesirable signal characteristics such as random noise is required. This can be done in a quite elegant and efficient way by the minimum description length (MDL) principle, which treats and separates ‘noise’ from the useful information as that part in the data that cannot be compressed. In its current form the MDL denoising method assumes the Gaussian noise model but does not require any ad hoc parameter settings. It provides a basis for high‐speed automated processing systems without requiring continual user interventions to validate the results as in the conventional signal processing methods. Our analysis of the denoising problem in mass spectrometry, capillary electrophoresis genotyping, and sequencing signals suggests that the MDL denoising method produces robust and intuitively appealing results sometimes even in situations where competing approaches perform poorly.

Monitoring therapy responses at the leukemic subclone level by ultra-deep amplicon resequencing in acute myeloid leukemia

In our individualized systems medicine program, personalized treatment options are identified and administered to chemorefractory acute myeloid leukemia (AML) patients based on exome sequencing and ex vivo drug sensitivity and resistance testing data. Here, we analyzed how clonal heterogeneity affects the responses of 13 AML patients to chemotherapy or targeted treatments using ultra-deep (average 68 000 × coverage) amplicon resequencing. Using amplicon resequencing, we identified 16 variants from 4 patients (frequency 0.54–2%) that were not detected previously by exome sequencing. A correlation-based method was developed to detect mutation-specific responses in serial samples across multiple time points. Significant subclone-specific responses were observed for both chemotherapy and targeted therapy. We detected subclonal responses in patients where clinical European LeukemiaNet (ELN) criteria showed no response. Subclonal responses also helped to identify putative mechanisms underlying drug sensitivities, such as sensitivity to azacitidine in DNMT3A mutated cell clones and resistance to cytarabine in a subclone with loss of NF1 gene. In summary, ultra-deep amplicon resequencing method enables sensitive quantification of subclonal variants and their responses to therapies. This approach provides new opportunities for designing combinatorial therapies blocking multiple subclones as well as for real-time assessment of such treatments.

Delivering ICT infrastructure for biomedical research

This paper describes an implementation of the Infrastructure-as-a-Service (IaaS) concept for scientific computing and seven service pilot implementations with requirements from biomedical use cases at the CSC - IT Center for Science. The key service design requirements were enabling the use of any scientific software environment the use cases needed to succeed, and delivering the distributed infrastructure ICT resources seamlessly with the local ICT resources for the scientist users. The service concept targets the IT administrators at research organisations and delivers virtualised compute cluster and storage capacity via private network solutions. The virtualised resources can become part of the local cluster as virtual nodes and they can share the same file system as the physical nodes assuming the network performance is sufficient. Extension of the local resources can then be made transparent to enable an easy infrastructure uptake to the scientist end-users. Based on 20 months of service piloting most of the biomedical organisations express a sustained and growing need for the distributed compute and storage resources delivered with the IaaS. We conclude that a successful implementation of the IaaS can improve access and reduce the effort to run expensive ICT infrastructure needed for biomedical research.

The Body Politic: Husserl and the Embodied Community

This article elucidates Edmund Husserl’s theory of community by examining its critical relation to the tradition of body politic, that is, the philosophical current employing the analogy between community and human body. It is argued that Husserl employs the corporeal analogy for three purposes: to describe the peculiar materiality, autonomy as well as the normative ideal of the social collective. Despite his relentless critique of naturalistic concepts in the description of the social sphere, Husserl nevertheless resorted to organic and bodily metaphors in his attempts to delineate the ideal form of communal co-existence, the “community of love”. With the help of the notion of love, Husserl was able to articulate his most elaborate account on the authentic relation between the individual and the collective.

Abstract 2378: Responses of AML patients to tailored drug regimens: monitoring cancer subclones by ultra-deep resequencing

As part of our individualized systems medicine (ISM) program, personalized treatment options are provided to clinicians based on in-depth genomic and molecular profiling as well as ex vivo drug sensitivity and resistance testing (DSRT) of leukemia patients (Pemovska et al. Cancer Discovery, 2013). In chemorefractory AML patients (n = 17), the ISM strategy has resulted in up to 35% response rate when individually selected targeted drugs have been applied in patient treatment. The responses achieved have, however, been transient and patients have typically relapsed quickly. Here, we aimed to understand the molecular basis of such treatment failures by quantitating the kinetics of individual cancer subclones before, during and after targeted treatments, as well as at the time of relapse and disease progression. Longitudinal serial samples from 13 AML patients were studied at multiple steps during leukemia progression and drug response. Clonal evolution of leukemic subclones was studied by both exome sequencing to get genome-wide overviews of disease progression, as well as by ultra-deep (>10,000x) amplicon resequencing with unique molecular identifiers to identify rare clones carrying specific cancer-relevant mutations. Nine of the 13 patients (69%) had multiple clones by exome sequencing and displayed branching evolution. In five patients who received treatment with targeted inhibitors we observed a significant differential therapeutic response of the individual AML subclones during therapy. In some cases, this could be directly attributed to the molecular mechanisms of drug response and resistance, such as the loss of NF1 in a subclone leading to cytarabine resistance or the loss of the FLT3-positive subclone in a patient responding to sunitinib treatment. Despite a prominent drug response at the level of the subclone carrying the driver mutations, in all these patients a new subclone emerged that led to progression of the disease. In three of the patients, the dominant clone appearing at relapse was already detected as a minor subclone in the diagnostic sample by amplicon resequencing. Amplicon sequencing enabled us to detect these minor subclones (down to 0,5% frequency) that were missed by exome sequencing. The results suggest that relapses in AML may arise because the drug-resistant subclone exists already before the onset of therapy. Overall, it is necessary to quantify tumor evolution and drug responses at the level of cancer subclones. Ultra-deep resequencing can be used to monitor drug responses at the subclone level, even at very low frequencies. This could facilitate early detection of small subclones with important prognostic implications, as well as the design of intelligent combinations of targeted drugs that could block such subclones. Citation Format: Poojitha N Ojamies, Mika Kontro, Henrik Edgren, Pekka Ellonen, Sonja Lagstrom, Henrikki Almusa, Timo Miettinen, Samuli Eldfors, David Tamborero, Krister Wennerberg, Caroline Heckman, Kimmo Porkka, Maija Wolf, Olli Kallioniemi. Responses of AML patients to tailored drug regimens: monitoring cancer subclones by ultra-deep resequencing. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2378.

The Particular Universal: Europe in Modern Philosophies of History

In contemporary debates, Europe is most often referred to as a geographical, cultural, historical, or political-economic entity. But is it possible to conceive of Europe also as a philosophical idea?

On the Philosophical Foundations of Universalism: Reason, Task, Critique

This article investigates the philosophical history of European universalism with the aim of differentiating between its two senses: the modern and the Ancient. Based on Edmund Husserl’s late interpretations on the unique character of Greek philosophy, this distinction is articulated in terms of “substantial” and “formal” accounts of universalism. Against the modern (substantial) idea of universalism, which took its point of departure especially from the natural law theories of the early modern period, Husserl conceived Greek universalism as an essentially formal notion, which relied on the critique of one’s cultural-historical situation on the basis of the shared faculty of reason. Instead of a ready-made position, this idea of universalism is best described in terms of a “task”, which has its peculiar temporal horizon in infinity. By discussing the political implications of philosophical universalism, the article aims to uncover its latent cultural implications, that is, the ideas of self-critique and self-renewal nurturing the utopian motive of culture. Thus by broadening the philosophical scope of universalism, the article will insist on its relevance for contemporary debate on