Correction: Whole-genome sequencing of chronic lymphocytic leukaemia reveals distinct differences in the mutational landscape between IgHVmut and IgHVunmut subgroups.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Whole-genome sequencing of chronic lymphocytic leukaemia reveals distinct differences in the mutational landscape between IgHVmut and IgHVunmut subgroups.

Chronic lymphocytic leukaemia (CLL) consists of two biologically and clinically distinct subtypes defined by the abundance of somatic hypermutation (SHM) affecting the Ig variable heavy-chain locus (IgHV). The molecular mechanisms underlying these subtypes are incompletely understood. Here, we present a comprehensive whole-genome sequencing analysis of somatically acquired genetic events from 46 CLL patients, including a systematic comparison of coding and non-coding single-nucleotide variants, copy number variants and structural variants, regions of kataegis and mutation signatures between IgHVmut and IgHVunmut subtypes. We demonstrate that one-quarter of non-coding mutations in regions of kataegis outside the Ig loci are located in genes relevant to CLL. We show that non-coding mutations in ATM may negatively impact on ATM expression and find non-coding and regulatory region mutations in TCL1A, and in IgHVunmut CLL in IKZF3, SAMHD1,PAX5 and BIRC3. Finally, we show that IgHVunmut CLL is dominated by coding mutations in driver genes and an aging signature, whereas IgHVmut CLL has a high incidence of promoter and enhancer mutations caused by aberrant activation-induced cytidine deaminase activity. Taken together, our data support the hypothesis that differences in clinical outcome and biological characteristics between the two subgroups might reflect differences in mutation distribution, incidence and distinct underlying mutagenic mechanisms.

Whole-genome sequencing of chronic lymphocytic leukaemia reveals distinct differences in the mutational landscape between IgHVmut and IgHVunmut subgroups.

This corrects the article DOI: 10.1038/leu.2017.177.

Targeted deep sequencing reveals clinically relevant subclonal IgHV rearrangements in chronic lymphocytic leukemia.

The immunoglobulin heavy-chain variable region gene (IgHV) mutational status is considered the gold standard of prognostication in chronic lymphocytic leukemia (CLL) and is currently determined by Sanger sequencing that allows the analysis of the major clone. Using next-generation sequencing (NGS), we sequenced the IgHV gene from two independent cohorts: (A) 270 consecutive patient samples obtained at diagnosis and (B) 227 patients from the UK ARCTIC-AdMIRe clinical trials. Using complementary DNA from purified CD19+CD5+ cells, we demonstrate the presence of multiple rearrangements in independent experiments and showed that 24.4% of CLL patients express multiple productive clonally unrelated IgHV rearrangements. On the basis of IgHV-NGS subclonal profiles, we defined five different categories: patients with (a) multiple hypermutated (M) clones, (b) 1 M clone, (c) a mix of M-unmutated (UM) clones, (d) 1 UM clone and (e) multiple UM clones. In population A, IgHV-NGS classification stratified patients into five different subgroups with median treatment-free survival (TFS) of >280(a), 131(b), 94(c), 29(d), 15(e) months (P<0.0001) and a median OS of >397(a), 292(b), 196(c), 137(d) and 100(e) months (P<0.0001). In population B, the poor prognosis of multiple UM patients was confirmed with a median TFS of 2 months (P=0.0038). In conclusion, IgHV-NGS highlighted one quarter of CLL patients with multiple productive IgHV subclones and improves disease stratification and raises important questions concerning the pre-leukemic cellular origin of CLL.

Quantification of subclonal distributions of recurrent genomic aberrations in paired pre-treatment and relapse samples from patients with B-cell chronic lymphocytic leukemia.

Genome-wide array approaches and sequencing analyses are powerful tools for identifying genetic aberrations in cancers, including leukemias and lymphomas. However, the clinical and biological significance of such aberrations and their subclonal distribution are poorly understood. Here, we present the first genome-wide array based study of pre-treatment and relapse samples from patients with B-cell chronic lymphocytic leukemia (B-CLL) that uses the computational statistical tool OncoSNP. We show that quantification of the proportion of copy number alterations (CNAs) and copy neutral loss of heterozygosity regions (cnLOHs) in each sample is feasible. Furthermore, we (i) reveal complex changes in the subclonal architecture of paired samples at relapse compared with pre-treatment, (ii) provide evidence supporting an association between increased genomic complexity and poor clinical outcome (iii) report previously undefined, recurrent CNA/cnLOH regions that expand or newly occur at relapse and therefore might harbor candidate driver genes of relapse and/or chemotherapy resistance. Our findings are likely to impact on future therapeutic strategies aimed towards selecting effective and individually tailored targeted therapies.

Haemoglobin (Hb) G-Philadelphia, Hb Stanleyville-II, Hb G-Norfolk, Hb Matsue-Oki and Hb Mizushi can form a panel of α-chain variants that overlap in their phenotype: the novel use of StyI to screen for Hb G-Philadelphia.

INTRODUCTION: Haemoglobin (Hb) G-Philadelphia mutation is a common alpha-globin chain variant [α68(E17)Asn > Lys]. Combined high performance liquid chromatography (HPLC) and isoelectric focusing (IEF) can be used in a presumptive diagnosis of Hb G-Philadelphia, but there are other α-chain variants with a similar phenotype that cannot be excluded. Our aim was to develop a novel StyI restriction enzyme assay to diagnose the common Hb G-Philadelphia mutation and to identify any other variants with a similar phenotype by DNA sequencing. METHODS: Thirty-one cases given a presumptive diagnosis as Hb G-Philadelphia by HPLC and IEF were subjected to DNA analysis by restriction enzyme digestion using StyI. Negative cases were then subjected to DNA sequencing. RESULTS: Twenty-two cases (78.6%) of 28 cases amplified were tested positive for Hb G-Philadelphia by StyI restriction digestion. Sequencing of the six negative cases revealed two cases of Hb G-Philadelphia with C→A mutation in codon 68 in α2 globin gene, plus one case each of Hb G-Norfolk Hb Stanleyville-II, Hb Matsue-Oki and Hb Mizushi. CONCLUSION: A novel StyI restriction enzyme can be used to confirm the commonest type of Hb G-Philadelphia. DNA sequencing identified four other α-chain variants with a similar HPLC and IEF phenotype.

Measurement of compliance of the maternal abdominal wall in pregnancy.

Recent importance in the signal-to-noise ratio of fetal phonocardiograms has been attributed to a 'compliance-matching' between the phono-sensor and the maternal abdominal wall. We have developed a device to measure objectively the compliance of the maternal abdominal wall in pregnancy. Using this we have shown that the compliance varies considerably from patient to patient and that it varies most significantly (and inversely) with gestational age. As a result of this study we are now in a better position to optimize the compliance-matching of phono-sensors and thereby to detect fetal heart sounds with much more fidelity than hitherto.

An experimental rig to simulate fetal heart sounds.

Renewed interest in fetal phonocardiography has brought about new phono-sensor designs. We have developed an experimental rig to simulate generation of fetal heart sounds and also their passage through to the maternal abdominal wall. Using this rig we have investigated the design techniques applied to the new phono-sensors and we have also analysed the sensors themselves. We can now approach an ideal sensor design for the faithful detection of fetal heart sounds.

Fatigue fractures of Thompson prostheses. A clinical and biomechanical study of revision procedures.

In four patients with fatigue fractures of Thompson prostheses, all had been in place for more than five years. All the patients were very active following the original operative procedure. Radiographs showed a varus position of the implant in three of the four patients, and the proximal fragment appeared loose; this was confirmed at revision operation. The distal fragment was rigidly held, and was only removed after the femoral shaft had been fenestrated. If a window had been cut in the shaft of the femur, the bone was considerably weakened. This clinical impression was confirmed by cadaver studies, which revealed that a lateral window weakens the femur more than an anterior window. For revision operations, replacement with a long-stemmed prosthesis is desirable. This protects the weakened segment of the femur, and provides sufficient strength for immediate weight-bearing.