Circulating tumour DNA (ctDNA) assays conducted on plasma are rapidly developing a strong evidence base for use in patients with cancer. The European Society for Medical Oncology convened an expert ...working group to review the analytical and clinical validity and utility of ctDNA assays. For patients with advanced cancer, validated and adequately sensitive ctDNA assays have utility in identifying actionable mutations to direct targeted therapy, and may be used in routine clinical practice, provided the limitations of the assays are taken into account. Tissue-based testing remains the preferred test for many cancer patients, due to limitations of ctDNA assays detecting fusion events and copy number changes, although ctDNA assays may be routinely used when faster results will be clinically important, or when tissue biopsies are not possible or inappropriate. Reflex tumour testing should be considered following a non-informative ctDNA result, due to false-negative results with ctDNA testing. In patients treated for early-stage cancers, detection of molecular residual disease or molecular relapse, has high evidence of clinical validity in anticipating future relapse in many cancers. Molecular residual disease/molecular relapse detection cannot be recommended in routine clinical practice, as currently there is no evidence for clinical utility in directing treatment. Additional potential applications of ctDNA assays, under research development and not recommended for routine practice, include identifying patients not responding to therapy with early dynamic changes in ctDNA levels, monitoring therapy for the development of resistance mutations before clinical progression, and in screening asymptomatic people for cancer. Recommendations for reporting of results, future development of ctDNA assays and future clinical research are made.
Several professional societies have published guidelines for the clinical interpretation of somatic variants, which specifically address diagnostic, prognostic, and therapeutic implications. Although ...these guidelines for the clinical interpretation of variants include data types that may be used to determine the oncogenicity of a variant (eg, population frequency, functional, and in silico data or somatic frequency), they do not provide a direct, systematic, and comprehensive set of standards and rules to classify the oncogenicity of a somatic variant. This insufficient guidance leads to inconsistent classification of rare somatic variants in cancer, generates variability in their clinical interpretation, and, importantly, affects patient care. Therefore, it is essential to address this unmet need.
Clinical Genome Resource (ClinGen) Somatic Cancer Clinical Domain Working Group and ClinGen Germline/Somatic Variant Subcommittee, the Cancer Genomics Consortium, and the Variant Interpretation for Cancer Consortium used a consensus approach to develop a standard operating procedure (SOP) for the classification of oncogenicity of somatic variants.
This comprehensive SOP has been developed to improve consistency in somatic variant classification and has been validated on 94 somatic variants in 10 common cancer-related genes.
The comprehensive SOP is now available for classification of oncogenicity of somatic variants.
Osteoarthritis (OA) is the most prevalent form of arthritis and accounts for substantial morbidity and disability, particularly in older people. It is characterised by changes in joint structure, ...including degeneration of the articular cartilage, and its aetiology is multifactorial with a strong postulated genetic component.
A meta-analysis was performed of four genome-wide association (GWA) studies of 2371 cases of knee OA and 35 909 controls in Caucasian populations. Replication of the top hits was attempted with data from 10 additional replication datasets.
With a cumulative sample size of 6709 cases and 44 439 controls, one genome-wide significant locus was identified on chromosome 7q22 for knee OA (rs4730250, p=9.2 × 10⁻⁹), thereby confirming its role as a susceptibility locus for OA.
The associated signal is located within a large (500 kb) linkage disequilibrium block that contains six genes: PRKAR2B (protein kinase, cAMP-dependent, regulatory, type II, β), HPB1 (HMG-box transcription factor 1), COG5 (component of oligomeric golgi complex 5), GPR22 (G protein-coupled receptor 22), DUS4L (dihydrouridine synthase 4-like) and BCAP29 (B cell receptor-associated protein 29). Gene expression analyses of the (six) genes in primary cells derived from different joint tissues confirmed expression of all the genes in the joint environment.
The double-strand break repair (DSBR) model of recombination predicts that heteroduplexes will be formed in regions that flank the double-strand break (DSB) site and that the resulting intermediate ...is resolved to generate either crossovers or noncrossovers for flanking markers. Previous studies in Saccharomyces cerevisiae, however, failed to detect heteroduplexes on both sides of the DSB site. Recent physical studies suggest that some recombination events involve heterodupex formation by a mechanism, synthesis-dependent strand annealing (SDSA), that is inherently asymmetric with respect to the DSB site and that leads exclusively to noncrossovers of flanking markers. Below, we demonstrate that many of the recombination events initiated at the HIS4 recombination hotspot are consistent with a variant of the DSBR model in which the extent of heteroduplex on one side of the DSB site is much greater than that on the other. Events that include only one flanking marker in the heteroduplex (unidirectional events) are usually resolved as noncrossovers, whereas events that include both flanking markers (bidirectional events) are usually resolved as crossovers. The unidirectional events may represent SDSA, consistent with the conclusions of others, although other possibilities are not excluded. We also show that the level of recombination reflects the integration of events initiated at several different DSB sites, and we identify a subset of gene conversion events that may involve break-induced replication (BIR) or repair of a double-stranded DNA gap.
In order to identify novel somatic mutations associated with classic BCR/ABL1-negative myeloproliferative neoplasms, we performed high-coverage genome sequencing of DNA from peripheral blood ...granulocytes and cultured skin fibroblasts from a patient with MPL W515K-positive primary myelofibrosis. The primary myelofibrosis genome had a low somatic mutation rate, consistent with that observed in similar hematopoietic tumor genomes. Interfacing of whole-genome DNA sequence data with RNA expression data identified three somatic mutations of potential functional significance: i) a nonsense mutation in CARD6, implicated in modulation of NF-kappaB activation; ii) a 19-base pair deletion involving a potential regulatory region in the 5'-untranslated region of BRD2, implicated in transcriptional regulation and cell cycle control; and iii) a non-synonymous point mutation in KIAA0355, an uncharacterized protein. Additional mutations in three genes (CAP2, SOX30, and MFRP) were also evident, albeit with no support for expression at the RNA level. Re-sequencing of these six genes in 178 patients with polycythemia vera, essential thrombocythemia, and myelofibrosis did not identify recurrent somatic mutations in these genes. Finally, we describe methods for reducing false-positive variant calls in the analysis of hematologic malignancies with a low somatic mutation rate. This trial is registered with ClinicalTrials.gov (NCT01108159).
Borde and colleagues reported that a reporter plasmid inserted at different genomic locations in Saccharomyces cerevisiae had different levels of meiotic recombination activity. We show that the ...level of recombination activity is very significantly correlated with the GC content of DNA sequences flanking the insertion.
This article provides recommendations for the care of laboratory zebrafish (Danio rerio) as part of the further implementation of Annex A to the European Convention on the protection of vertebrate ...animals used for experimental and other scientific purposes, EU Commission Recommendation 2007/526/EC and the fulfilment of Article 33 of EU Directive 2010/63, both concerning the housing and care of experimental animals. The recommendations provide guidance on best practices and ranges of husbandry parameters within which zebrafish welfare, as well as reproducibility of experimental procedures, are assured.
Husbandry procedures found today in zebrafish facilities are numerous. While the vast majority of these practices are perfectly acceptable in terms of zebrafish physiology and welfare, the reproducibility of experimental results could be improved by further standardisation of husbandry procedures and exchange of husbandry information between laboratories. Standardisation protocols providing ranges of husbandry parameters are likely to be more successful and appropriate than the implementation of a set of fixed guidance values neglecting the empirically successful daily routines of many facilities and will better reflect the wide range of environmental parameters that characterise the natural habitats occupied by zebrafish.
A joint working group on zebrafish housing and husbandry recommendations, with members of the European Society for Fish Models in Biology and Medicine (EUFishBioMed) and of the Federation of European Laboratory Animal Science Associations (FELASA) has been given a mandate to provide guidelines based on a FELASA list of parameters, ‘Terms of Reference’.
The rate of meiotic recombination in the yeast Saccharomyces cerevisiae varies widely in different regions of the genome with some genes having very high levels of recombination (hotspots). A variety ...of experiments done in yeast suggest that hotspots are a feature of chromatin structure rather than a feature of primary DNA sequence. We examined the effects of mutating a variety of enzymes that affect chromatin structure on the recombination activity of the well-characterized HIS4 hotspot including the Set2p and Dot1p histone methylases, the Hda1p and Rpd3p histone deacetylases, the Sin4p global transcription regulator, and a deletion of one of the two copies of the genes encoding histone H3–H4. Loss of Set2p or Rpd3p substantially elevated HIS4 hotspot activity, and loss of Hda1p had a smaller stimulatory effect; none of the other alterations had a significant effect. The increase of HIS4 hotspot activity in set2 and rpd3 strains is likely to be related to the recent finding that histone H3 methylation by Set2p directs deacetylation of histones by Rpd3p.
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