199468814
Mutational processes contributing to the development of multiple myeloma
To gain insight into multiple myeloma (MM) tumorigenesis, we analyzed the mutational signatures in 874 whole-exome and 850 whole-genome data from the CoMMpass Study. We identified that coding and non-coding regions are differentially dominated by distinct single-nucleotide variant (SNV) mutational signatures, as well as five de novo structural rearrangement signatures. Mutational signatures reflective of different principle mutational processes—aging, defective DNA repair, and apolipoprotein B editing complex (APOBEC)/activation-induced deaminase activity—characterize MM. These mutational signatures show evidence of subgroup specificity—APOBEC-attributed signatures associated with MAF translocation t(14;16) and t(14;20) MM; potentially DNA repair deficiency with t(11;14) and t(4;14); and aging with hyperdiploidy. Mutational signatures beyond that associated with APOBEC are independent of established prognostic markers and appear to have relevance to predicting high-risk MM.
3912427
Signatures of mutational processes in human cancer
All cancers are caused by somatic mutations; however, understanding of the biological processes generating these mutations is limited. The catalogue of somatic mutations from a cancer genome bears the signatures of the mutational processes that have been operative. Here we analysed 4,938,362 mutations from 7,042 cancers and extracted more than 20 distinct mutational signatures. Some are present in many cancer types, notably a signature attributed to the APOBEC family of cytidine deaminases, whereas others are confined to a single cancer class. Certain signatures are associated with age of the patient at cancer diagnosis, known mutagenic exposures or defects in DNA maintenance, but many are of cryptic origin. In addition to these genome-wide mutational signatures, hypermutation localized to small genomic regions, ‘kataegis’, is found in many cancer types. The results reveal the diversity of mutational processes underlying the development of cancer, with potential implications for understanding of cancer aetiology, prevention and therapy.
10931544
Mechanisms underlying mutational signatures in human cancers
The collective somatic mutations observed in a cancer are the outcome of multiple mutagenic processes that have been operative over the lifetime of a patient. Each process leaves a characteristic imprint — a mutational signature — on the cancer genome, which is defined by the type of DNA damage and DNA repair processes that result in base substitutions, insertions and deletions or structural variations. With the advent of whole-genome sequencing, researchers are identifying an increasing array of these signatures. Mutational signatures can be used as a physiological readout of the biological history of a cancer and also have potential use for discerning ongoing mutational processes from historical ones, thus possibly revealing new targets for anticancer therapies.
13927527
Deciphering Signatures of Mutational Processes Operative in Human Cancer
Summary The genome of a cancer cell carries somatic mutations that are the cumulative consequences of the DNA damage and repair processes operative during the cellular lineage between the fertilized egg and the cancer cell. Remarkably, these mutational processes are poorly characterized. Global sequencing initiatives are yielding catalogs of somatic mutations from thousands of cancers, thus providing the unique opportunity to decipher the signatures of mutational processes operative in human cancer. However, until now there have been no theoretical models describing the signatures of mutational processes operative in cancer genomes and no systematic computational approaches are available to decipher these mutational signatures. Here, by modeling mutational processes as a blind source separation problem, we introduce a computational framework that effectively addresses these questions. Our approach provides a basis for characterizing mutational signatures from cancer-derived somatic mutational catalogs, paving the way to insights into the pathogenetic mechanism underlying all cancers.
196664064
Mutational signatures associated with tobacco smoking in human cancer
Tobacco smoking increases the risk of at least 15 classes of cancer. We analyzed somatic mutations and DNA methylation in 5,243 cancers of types for which tobacco smoking confers an elevated risk. Smoking is associated with increased mutation burdens of multiple distinct mutational signatures, which contribute to different extents in different cancers. One of these signatures, mainly found in cancers derived from tissues directly exposed to tobacco smoke, is attributable to misreplication of DNA damage caused by tobacco carcinogens. Others likely reflect indirect activation of DNA editing by APOBEC cytidine deaminases and of an endogenous clock-like mutational process. The results are consistent with the proposition that smoking increases cancer risk by increasing the somatic mutation load, although direct evidence for this mechanism is lacking in some smoking-related cancer types. ONE SENTENCE SUMMARY Multiple distinct mutational processes associate with tobacco smoking in cancer reflecting direct and indirect effects of tobacco smoke.
779646
Environmental exposures and mutational patterns of cancer genomes
The etiology of most human cancers is unknown. Genetic inheritance and environmental factors are thought to have major roles, and for some types of cancer, exposure to carcinogens is a proven mechanism leading to tumorigenesis. Sequencing of entire cancer genomes has not only begun to provide clues regarding functionally relevant mutations, but has also paved the way towards understanding the initial exposures leading to DNA damage, repair and eventually to mutation of specific sequences within a cancer genome. Two recent studies of melanoma and small cell lung cancer exemplify what type of information can be gained from cancer genome sequencing.
13555473
Genomic complexity of multiple myeloma and its clinical implications
Multiple myeloma (MM) is a genetically complex disease that evolves from pre-malignant stages, such as monoclonal gammaopathy of undetermined significance and smouldering multiple myeloma, and progresses to symptomatic MM; this continuum provides a unique framework to study the sequential genomic evolution of MM. In the past 5 years, results from large-scale whole-exome sequencing studies have provided new insights into the clonal heterogeneity and evolution of the disease. Moreover, the recurrent co-occurrence of genomic events helps to dissect the genomic complexity underlying tumour progression. According to the primary genetic events involved in tumorigenesis, MM tumours are hierarchically subdivided into hyperdiploid and non-hyperdiploid subtypes; subsequently, secondary genetic events lead to tumour progression. In this Review, we describe the 'driver' gene alterations involved in the development and progression of MM, with a focus on the sequential acquisition of the main genomic aberrations. We also provide valuable insight into the clonal heterogeneity and clonal evolution of the disease, as well as into the therapeutic implications of a comprehensive understanding of the genomic complexity of MM.
30711359
Mutational Spectrum, Copy Number Changes, and Outcome: Results of a Sequencing Study of Patients With Newly Diagnosed Myeloma.
PURPOSE
At the molecular level, myeloma is characterized by copy number abnormalities and recurrent translocations into the immunoglobulin heavy chain locus. Novel methods, such as massively parallel sequencing, have begun to describe the pattern of tumor-acquired mutations, but their clinical relevance has yet to be established.


METHODS
We performed whole-exome sequencing for 463 patients who presented with myeloma and were enrolled onto the National Cancer Research Institute Myeloma XI trial, for whom complete molecular cytogenetic and clinical outcome data were available.


RESULTS
We identified 15 significantly mutated genes: IRF4, KRAS, NRAS, MAX, HIST1H1E, RB1, EGR1, TP53, TRAF3, FAM46C, DIS3, BRAF, LTB, CYLD, and FGFR3. The mutational spectrum is dominated by mutations in the RAS (43%) and nuclear factor-κB (17%) pathways, but although they are prognostically neutral, they could be targeted therapeutically. Mutations in CCND1 and DNA repair pathway alterations (TP53, ATM, ATR, and ZNFHX4 mutations) are associated with a negative impact on survival. In contrast, those in IRF4 and EGR1 are associated with a favorable overall survival. We combined these novel mutation risk factors with the recurrent molecular adverse features and international staging system to generate an international staging system mutation score that can identify a high-risk population of patients who experience relapse and die prematurely.


CONCLUSION
We have refined our understanding of genetic events in myeloma and identified clinically relevant mutations that may be used to better stratify patients at presentation.
7318613
Heterogeneity of genomic evolution and mutational profiles in multiple myeloma
Multiple myeloma is an incurable plasma cell malignancy with a complex and incompletely understood molecular pathogenesis. Here we use whole-exome sequencing, copy-number profiling and cytogenetics to analyse 84 myeloma samples. Most cases have a complex subclonal structure and show clusters of subclonal variants, including subclonal driver mutations. Serial sampling reveals diverse patterns of clonal evolution, including linear evolution, differential clonal response and branching evolution. Diverse processes contribute to the mutational repertoire, including kataegis and somatic hypermutation, and their relative contribution changes over time. We find heterogeneity of mutational spectrum across samples, with few recurrent genes. We identify new candidate genes, including truncations of SP140, LTB, ROBO1 and clustered missense mutations in EGR1. The myeloma genome is heterogeneous across the cohort, and exhibits diversity in clonal admixture and in dynamics of evolution, which may impact prognostic stratification, therapeutic approaches and assessment of disease response to treatment.
206541703
Widespread genetic heterogeneity in multiple myeloma: implications for targeted therapy.
We performed massively parallel sequencing of paired tumor/normal samples from 203 multiple myeloma (MM) patients and identified significantly mutated genes and copy number alterations and discovered putative tumor suppressor genes by determining homozygous deletions and loss of heterozygosity. We observed frequent mutations in KRAS (particularly in previously treated patients), NRAS, BRAF, FAM46C, TP53, and DIS3 (particularly in nonhyperdiploid MM). Mutations were often present in subclonal populations, and multiple mutations within the same pathway (e.g., KRAS, NRAS, and BRAF) were observed in the same patient. In vitro modeling predicts only partial treatment efficacy of targeting subclonal mutations, and even growth promotion of nonmutated subclones in some cases. These results emphasize the importance of heterogeneity analysis for treatment decisions.
7590999
Mutational signatures reveal the dynamic interplay of risk factors and cellular processes during liver tumorigenesis
Genomic alterations driving tumorigenesis result from the interaction of environmental exposures and endogenous cellular processes. With a diversity of risk factors, liver cancer is an ideal model to study these interactions. Here, we analyze the whole genomes of 44 new and 264 published liver cancers and we identify 10 mutational and 6 structural rearrangement signatures showing distinct relationships with environmental exposures, replication, transcription, and driver genes. The liver cancer-specific signature 16, associated with alcohol, displays a unique feature of transcription-coupled damage and is the main source of CTNNB1 mutations. Flood of insertions/deletions (indels) are identified in very highly expressed hepato-specific genes, likely resulting from replication-transcription collisions. Reconstruction of sub-clonal architecture reveals mutational signature evolution during tumor development exemplified by the vanishing of aflatoxin B1 signature in African migrants. Finally, chromosome duplications occur late and may represent rate-limiting events in tumorigenesis. These findings shed new light on the natural history of liver cancers.Tumorigenesis is a complex process driven by numerous risk factors. Here, genomic analysis of liver cancer reveals the evolution of mutational signatures during tumor development, highlighting mutational and structural signatures linked to environmental exposures and endogenous cellular processes.
8314066
Mutational Processes Molding the Genomes of 21 Breast Cancers
Summary All cancers carry somatic mutations. The patterns of mutation in cancer genomes reflect the DNA damage and repair processes to which cancer cells and their precursors have been exposed. To explore these mechanisms further, we generated catalogs of somatic mutation from 21 breast cancers and applied mathematical methods to extract mutational signatures of the underlying processes. Multiple distinct single- and double-nucleotide substitution signatures were discernible. Cancers with BRCA1 or BRCA2 mutations exhibited a characteristic combination of substitution mutation signatures and a distinctive profile of deletions. Complex relationships between somatic mutation prevalence and transcription were detected. A remarkable phenomenon of localized hypermutation, termed “kataegis,” was observed. Regions of kataegis differed between cancers but usually colocalized with somatic rearrangements. Base substitutions in these regions were almost exclusively of cytosine at TpC dinucleotides. The mechanisms underlying most of these mutational signatures are unknown. However, a role for the APOBEC family of cytidine deaminases is proposed. PaperClip
13757169
Validating the concept of mutational signatures with isogenic cell models
The diversity of somatic mutations in human cancers can be decomposed into individual mutational signatures, patterns of mutagenesis that arise because of DNA damage and DNA repair processes that have occurred in cells as they evolved towards malignancy. Correlations between mutational signatures and environmental exposures, enzymatic activities and genetic defects have been described, but human cancers are not ideal experimental systems—the exposures to different mutational processes in a patient’s lifetime are uncontrolled and any relationships observed can only be described as an association. Here, we demonstrate the proof-of-principle that it is possible to recreate cancer mutational signatures in vitro using CRISPR-Cas9-based gene-editing experiments in an isogenic human-cell system. We provide experimental and algorithmic methods to discover mutational signatures generated under highly experimentally-controlled conditions. Our in vitro findings strikingly recapitulate in vivo observations of cancer data, fundamentally validating the concept of (particularly) endogenously-arising mutational signatures.As cells evolve towards malignancy, somatic mutations arise from defects in DNA damage and repair processes which are each associated with individual mutation signatures. Here the authors show it is possible to recreate cancer mutational signatures in vitro using gene editing experiments in an isogenic human-cell system.
2122093
Somatic ERCC2 Mutations Are Associated with a Distinct Genomic Signature in Urothelial Tumors
Alterations in DNA repair pathways are common in tumors and can result in characteristic mutational signatures; however, a specific mutational signature associated with somatic alterations in the nucleotide- excision repair (NER) pathway has not yet been identified. Here we examine the mutational processes operating in urothelial cancer, a tumor type in which the core NER gene ERCC2 is significantly mutated. Analysis of three independent urothelial tumor cohorts demonstrates a strong association between somatic ERCC2 mutations and the activity of a mutational signature characterized by a broad spectrum of base changes. In addition, we note an association between the activity of this signature and smoking that is independent of ERCC2 mutation status, providing genomic evidence of tobacco-related mutagenesis in urothelial cancer. Together, these analyses identify an NER-related mutational signature and highlight the related roles of DNA damage and subsequent DNA repair in shaping tumor mutational landscape.
195191529
Deficiency of nucleotide excision repair is associated with mutational signature observed in cancer.
Nucleotide excision repair (NER) is one of the main DNA repair pathways that protect cells against genomic damage. Disruption of this pathway can contribute to the development of cancer and accelerate aging. Mutational characteristics of NER-deficiency may reveal important diagnostic opportunities, as tumors deficient in NER are more sensitive to certain treatments. Here, we analyzed the genome-wide somatic mutational profiles of adult stem cells (ASCs) from NER-deficient Ercc1 -/Δ mice. Our results indicate that NER-deficiency increases the base substitution load twofold in liver but not in small intestinal ASCs, which coincides with the tissue-specific aging pathology observed in these mice. Moreover, NER-deficient ASCs of both tissues show an increased contribution of Signature 8 mutations, which is a mutational pattern with unknown etiology that is recurrently observed in various cancer types. The scattered genomic distribution of the base substitutions indicates that deficiency of global-genome NER (GG-NER) underlies the observed mutational consequences. In line with this, we observe increased Signature 8 mutations in a GG-NER-deficient human organoid culture, in which XPC was deleted using CRISPR-Cas9 gene-editing. Furthermore, genomes of NER-deficient breast tumors show an increased contribution of Signature 8 mutations compared with NER-proficient tumors. Elevated levels of Signature 8 mutations could therefore contribute to a predictor of NER-deficiency based on a patient's mutational profile.
7207558
Use of CRISPR-modified human stem cell organoids to study the origin of mutational signatures in cancer
A signature event for organoids Human cancer genomes harbor cryptic mutational signatures that represent the cumulative effects of DNA damage and defects in DNA repair processes. Knowledge of how specific signatures originate could have a major impact on cancer diagnosis and prevention. One approach to address this question is to reproduce the signatures in experimental systems by genetic engineering and then match the signatures to those found in naturally occurring cancers. Drost et al. used CRISPR-Cas9 to delete certain DNA repair enzymes from human colon organoids. In a proof-of-concept study, they show that deficiency in base excision repair is responsible for a mutational signature previously identified in cancer genome sequencing projects. Science, this issue p. 234 CRISPR-mediated genome editing of organoids can reveal the origin of mutational signatures in human cancer. Mutational processes underlie cancer initiation and progression. Signatures of these processes in cancer genomes may explain cancer etiology and could hold diagnostic and prognostic value. We developed a strategy that can be used to explore the origin of cancer-associated mutational signatures. We used CRISPR-Cas9 technology to delete key DNA repair genes in human colon organoids, followed by delayed subcloning and whole-genome sequencing. We found that mutation accumulation in organoids deficient in the mismatch repair gene MLH1 is driven by replication errors and accurately models the mutation profiles observed in mismatch repair–deficient colorectal cancers. Application of this strategy to the cancer predisposition gene NTHL1, which encodes a base excision repair protein, revealed a mutational footprint (signature 30) previously observed in a breast cancer cohort. We show that signature 30 can arise from germline NTHL1 mutations.
40573878
Molecular mechanisms of antibody somatic hypermutation.
Functional antibody genes are assembled by V-D-J joining and then diversified by somatic hypermutation. This hypermutation results from stepwise incorporation of single nucleotide substitutions into the V gene, underpinning much of antibody diversity and affinity maturation. Hypermutation is triggered by activation-induced deaminase (AID), an enzyme which catalyzes targeted deamination of deoxycytidine residues in DNA. The pathways used for processing the AID-generated U:G lesions determine the variety of base substitutions observed during somatic hypermutation. Thus, DNA replication across the uracil yields transition mutations at C:G pairs, whereas uracil excision by UNG uracil-DNA glycosylase creates abasic sites that can also yield transversions. Recognition of the U:G mismatch by MSH2/MSH6 triggers a mutagenic patch repair in which polymerase eta plays a major role and leads to mutations at A:T pairs. AID-triggered DNA deamination also underpins immunoglobulin variable (IgV) gene conversion, isotype class switching, and some oncogenic translocations in B cell tumors.
4451126
Non-coding recurrent mutations in chronic lymphocytic leukaemia
Chronic lymphocytic leukaemia (CLL) is a frequent disease in which the genetic alterations determining the clinicobiological behaviour are not fully understood. Here we describe a comprehensive evaluation of the genomic landscape of 452 CLL cases and 54 patients with monoclonal B-lymphocytosis, a precursor disorder. We extend the number of CLL driver alterations, including changes in ZNF292, ZMYM3, ARID1A and PTPN11. We also identify novel recurrent mutations in non-coding regions, including the 3′ region of NOTCH1, which cause aberrant splicing events, increase NOTCH1 activity and result in a more aggressive disease. In addition, mutations in an enhancer located on chromosome 9p13 result in reduced expression of the B-cell-specific transcription factor PAX5. The accumulative number of driver alterations (0 to ≥4) discriminated between patients with differences in clinical behaviour. This study provides an integrated portrait of the CLL genomic landscape, identifies new recurrent driver mutations of the disease, and suggests clinical interventions that may improve the management of this neoplasia.
86594213
Optimizing Cancer Genome Sequencing and Analysis
Tumors are typically sequenced to depths of 75-100× (exome) or 30-50× (whole genome). We demonstrate that current sequencing paradigms are inadequate for tumors that are impure, aneuploid or clonally heterogeneous. To reassess optimal sequencing strategies, we performed ultra-deep (up to ~312×) whole genome sequencing (WGS) and exome capture (up to ~433×) of a primary acute myeloid leukemia, its subsequent relapse, and a matched normal skin sample. We tested multiple alignment and variant calling algorithms and validated ~200,000 putative SNVs by sequencing them to depths of ~1,000×. Additional targeted sequencing provided over 10,000× coverage and ddPCR assays provided up to ~250,000× sampling of selected sites. We evaluated the effects of different library generation approaches, depth of sequencing, and analysis strategies on the ability to effectively characterize a complex tumor. This dataset, representing the most comprehensively sequenced tumor described to date, will serve as an invaluable community resource (dbGaP accession id phs000159).
4327775
Biological and prognostic impact of APOBEC-induced mutations in the spectrum of plasma cell dyscrasias and multiple myeloma cell lines
Biological and prognostic impact of APOBEC-induced mutations in the spectrum of plasma cell dyscrasias and multiple myeloma cell lines
5653455
deconstructSigs: delineating mutational processes in single tumors distinguishes DNA repair deficiencies and patterns of carcinoma evolution
BackgroundAnalysis of somatic mutations provides insight into the mutational processes that have shaped the cancer genome, but such analysis currently requires large cohorts. We develop deconstructSigs, which allows the identification of mutational signatures within a single tumor sample.ResultsApplication of deconstructSigs identifies samples with DNA repair deficiencies and reveals distinct and dynamic mutational processes molding the cancer genome in esophageal adenocarcinoma compared to squamous cell carcinomas.ConclusionsdeconstructSigs confers the ability to define mutational processes driven by environmental exposures, DNA repair abnormalities, and mutagenic processes in individual tumors with implications for precision cancer medicine.
8689666
Mutational Strand Asymmetries in Cancer Genomes Reveal Mechanisms of DNA Damage and Repair
Mutational processes constantly shape the somatic genome, leading to immunity, aging, cancer, and other diseases. When cancer is the outcome, we are afforded a glimpse into these processes by the clonal expansion of the malignant cell. Here, we characterize a less explored layer of the mutational landscape of cancer: mutational asymmetries between the two DNA strands. Analyzing whole-genome sequences of 590 tumors from 14 different cancer types, we reveal widespread asymmetries across mutagenic processes, with transcriptional ("T-class") asymmetry dominating UV-, smoking-, and liver-cancer-associated mutations and replicative ("R-class") asymmetry dominating POLE-, APOBEC-, and MSI-associated mutations. We report a striking phenomenon of transcription-coupled damage (TCD) on the non-transcribed DNA strand and provide evidence that APOBEC mutagenesis occurs on the lagging-strand template during DNA replication. As more genomes are sequenced, studying and classifying their asymmetries will illuminate the underlying biological mechanisms of DNA damage and repair.
7679324
Genomic evidence for elevated mutation rates in highly expressed genes
Reporter gene assays have demonstrated both transcription‐associated mutagenesis (TAM) and transcription‐coupled repair, but the net impact of transcription on mutation rate remains unclear, especially at the genomic scale. Using comparative genomics of related species as well as mutation accumulation lines, we show in yeast that the rate of point mutation in a gene increases with the expression level of the gene. Transcription induces mutagenesis on both DNA strands, indicating simultaneous actions of several TAM mechanisms. A significant positive correlation is also detected between the human germline mutation rate and expression level. These results indicate that transcription is overall mutagenic.
90836164
Analysis of Mutational Signatures Suggest That Aid Has an Early and Driver Role in Multiple Myeloma
INTRODUCTION: Multiple myeloma (MM) is a biologically and clinically heterogeneous disease. Different recurrent driver genomic events have been reported, but to date no unifying feature has been identified in MM evolution. The recent interest in signatures of mutational processes through analysis of whole-exome sequencing data has led to initial insights into what generates MM mutational repertoire (Bolli et al, Nat Com 2014). Here, taking advantage of the increased power provided by whole genome sequencing (WGS), we analyzed 22 paired samples from 11 patients first at the smoldering (SMM)/MGUS stage and subsequently at the time of progression to symptomatic MM to gain a deeper understanding of the full landscape of mutational processes operative in MM, especially during their evolution over time. MATERIAL AND METHODS: DNA from bone marrow CD138+ cells underwent WGS along with a matched normal sample using HiSeq X Ten machines (Illumina, Inc.). Mutational signature extraction was performed running non-negative matrix factorization (NMF) as previously described (Alexandrov et al, Nature 2013). RESULTS: We have observed and utilized a median number of 5780 (range 2599-7760) substitutions per patient at the asymptomatic stage and 5954 (ranges 2824-8227) at progression to MM stage to extract mutational signatures. NMF extracted 5 main signatures (http://cancer.sanger.ac.uk/cosmic/signatures). Specifically, APOBEC- (signature #2) and age-related signatures (signatures #1 and #5) accounted for 13% (1-21%) and 23% (3.2-40%) of all substitutions, respectively. In addition, we found two known signatures that were not implicated in MM so far: non-canonical AID (Signature #9), contributing to 28% of all substitutions (17-55%); and signature #8, accounting for 28% of all substitutions (13-45%) and pertaining to a yet unknown mutational process. Finally, the fifth signature did not match any of the previously described ones, representing a potential novel process which we defined as MM-1 (7%, range (1-16%). Interestingly, we found a differential contribution of processes in non-coding and intronic regions where AID was more prevalent, while exonic regions where APOBEC and age signatures were more prevalent. In intronic regions we found widespread regions of kataegis (9/11 patients), reflective of localized hypermutation. In our patients, kataegis was associated with rearrangements in 60% of cases, and was present in both the SMM and MM sample in 84% of cases, suggestive of an early event during tumor development. Contrary to what is observed in solid cancers, APOBEC signature was only responsible for 25% of kataegis variants, vs 70% for AID, suggesting a causative role of aberrant AID activity in shaping the early mutational repertoire of neoplastic plasma cells. To confirm this, we looked at serial samples in our cohort. While the percent contribution of each signature varied in each patient, confirming genomic heterogeneity of MM, it did not change when paired SMM and MM samples from the same patient were compared. This shows that mutational processes required for the development of symptomatic MM act early, and have been already operative at the SMM stage. However, by clustering substitutions as clonal (early variants present at the time of tumor initiation) or subclonal (late variants arisen closer to the time of sampling) using a Bayesian hierarchical Dirichlet process (Bolli et al, Nature Comms 2014), we could analyze processes operative before SMM was diagnosed. NMF analysis of these clusters reported striking differences. Specifically, AID and age were the predominant mutational processes in early substitutions in all patients, contributing to a median of 35% (25-54%) and 30% (15-43%) of variants respectively. Conversely the contribution of AID was minimal among late substitutions (5%, 1-22%), where instead APOBEC, Signature #8 and MM-1 activity was prominent [19% (1-43), 38% (8-73%), 16% (2-50%) respectively]. CONCLUSION: WGS data allowed the identification of mutational processes operative well before MM becomes clinically evident. Our observation that all samples have signs of aberrant AID at the time of tumor initiation supports a unifying model where MM precursors are initially transformed with the contribution of AID, providing a fertile ground for other later processes (i.e APOBEC and signature #8) to act and shape the final genomic landscape of overt multiple myeloma. Disclosures No relevant conflicts of interest to declare.
2284991
Landscape of somatic mutations in 560 breast cancer whole genome sequences
We analysed whole genome sequences of 560 breast cancers to advance understanding of the driver mutations conferring clonal advantage and the mutational processes generating somatic mutations. 93 protein-coding cancer genes carried likely driver mutations. Some non-coding regions exhibited high mutation frequencies but most have distinctive structural features probably causing elevated mutation rates and do not harbour driver mutations. Mutational signature analysis was extended to genome rearrangements and revealed 12 base substitution and six rearrangement signatures. Three rearrangement signatures, characterised by tandem duplications or deletions, appear associated with defective homologous recombination based DNA repair: one with deficient BRCA1 function; another with deficient BRCA1 or BRCA2 function; the cause of the third is unknown. This analysis of all classes of somatic mutation across exons, introns and intergenic regions highlights the repertoire of cancer genes and mutational processes operative, and progresses towards a comprehensive account of the somatic genetic basis of breast cancer.
5764656
The molecular classification of multiple myeloma.
To better define the molecular basis of multiple myeloma (MM), we performed unsupervised hierarchic clustering of mRNA expression profiles in CD138-enriched plasma cells from 414 newly diagnosed patients who went on to receive high-dose therapy and tandem stem cell transplants. Seven disease subtypes were validated that were strongly influenced by known genetic lesions, such as c-MAF- and MAFB-, CCND1- and CCND3-, and MMSET-activating translocations and hyperdiploidy. Indicative of the deregulation of common pathways by gene orthologs, common gene signatures were observed in cases with c-MAF and MAFB activation and CCND1 and CCND3 activation, the latter consisting of 2 subgroups, one characterized by expression of the early B-cell markers CD20 and PAX5. A low incidence of focal bone disease distinguished one and increased expression of proliferation-associated genes of another novel subgroup. Comprising varying fractions of each of the other 6 subgroups, the proliferation subgroup dominated at relapse, suggesting that this signature is linked to disease progression. Proliferation and MMSET-spike groups were characterized by significant overexpression of genes mapping to chromosome 1q, and both exhibited a poor prognosis relative to the other groups. A subset of cases with a predominating myeloid gene expression signature, excluded from the profiling analyses, had more favorable baseline characteristics and superior prognosis to those lacking this signature.
208548026
Apobec Family Mutational Signatures Are Associated with Poor Prognosis Translocations in Multiple Myeloma
Aberrant chromosomal translocations are seen in ~40% of presenting patients and predominantly involve the IGH locus at 14q32. The five main translocations involving the IGH locus are t(4;14), t(6;14), t(11;14), t(14;16) and t(14;20), which result in over-expression of MMSET/FGFR3, CCND3, CCND1, MAF and MAFB, respectively. In previous clinical trials we have shown that the t(4;14), t(14;16) and t(14;20) are associated with a poor prognosis. In initial sequencing studies of myeloma it has been noted that the spectrum of mutations fall into two groups, one of which is characterised by an APOBEC signature. This signature comprises of C>T, C>G and C>A mutations in a TpC context and comprises only a subset of samples, with the rest having a rather generic mutation signature representing an intrinsic mutational process occurring as a result of the spontaneous deamination of methylated cytosine to thymine. Whole exome sequencing was performed on 463 presentation patients enrolled into the UK Myeloma XI trial. DNA was extracted from germline DNA and CD138+ plasma cells and whole exome sequencing was performed using SureSelect (Agilent). In addition to capturing the exome, extra baits were added covering the IGH, IGK, IGL and MYCloci in order to determine the breakpoints associated with translocations in these genes. Tumor and germline DNA were sequenced to a median of 60x and data processed to generate copy number, acquired variants and translocation breakpoints in the tumor. Progression-free and overall survival was measured from initial randomization and median follow up for this analysis was 25 months. These combined data allow us to examine the effect of translocations on the mutational spectra in myeloma and determine any associations with progression-free or overall survival. Translocations were detected in 232 (50.1%) patients of which 59 patients (12.7%) had a t(4;14), 86 patients (18.6%) a t(11;14), 17 patients (3.7%) a t(14;16), 5 patients (1%) a t(6;14) and 4 patients (0.9%) a t(14;20). MYC translocations were found in 85 patients (18.4%). Using the tiled regions we were able to detect a mutational signature, kataegis, where regional clustering of mutations can be indicative of somatic genomic rearrangements. We found the hallmarks of kataegis in 15 samples (3.2%), where there was enrichment for TpCpH mutations with an inter-mutational distance We found mutation of translocation partner oncogenes, in particular CCND1 was mutated in 10 patients with the t(11;14). There was an association of mutated CCND1 with a poor prognosis when compared with non-mutated t(11;14) patients (OS median of 20.2 months vs. not reached, p=0.005). Mutations were also seen in FGFR3, MAF and MAFB but only in the samples with the respective translocations. The mutations are likely due to somatic hypermutation mediated by AID, an APOBEC family member. We found that t(14;16) and t(14;20) samples have a significantly higher number of mutations compare to the other translocation groups (p=1.65x10-5). These mutations were enriched for those with an APOBEC signature (T(C>T)A, p=9.1x10-5; T(C>T)T, p=0.0014; T(C>G)A, p=0.001; T(C>G)T, p=0.0064), indicating that the ‘maf’ translocation groups are characterized by APOBEC signature mutations, specifically APOBEC3B. When samples are assigned to either an APOBEC or non-APOBEC group the ‘maf’ translocations account for 66.6% of samples in the APOBEC group but only 1.3% of the non-APOBEC group. Here we show three different mutational signatures mediated by the APOBEC family: translocation partner mutation by AID, APOBEC signature mediated by APOBEC3B, and kataegis mediated by an unknown APOBEC family member. We also show for the first time a clinical impact of APOBEC mutations and their association with a poor prognosis. The poor prognosis of this mutational signature is inextricably linked to a high mutation load and the adverse t(14;16) and t(14;20) translocation subgroups. Disclosures Walker:Onyx Pharmaceuticals: Consultancy, Honoraria.
205352154
Human mutation rate associated with DNA replication timing
Eukaryotic DNA replication is highly stratified, with different genomic regions shown to replicate at characteristic times during S phase. Here we observe that mutation rate, as reflected in recent evolutionary divergence and human nucleotide diversity, is markedly increased in later-replicating regions of the human genome. All classes of substitutions are affected, suggesting a generalized mechanism involving replication time-dependent DNA damage. This correlation between mutation rate and regionally stratified replication timing may have substantial evolutionary implications.
2481405
How does DNA break during chromosomal translocations?
Chromosomal translocations are one of the most common types of genetic rearrangements and are molecular signatures for many types of cancers. They are considered as primary causes for cancers, especially lymphoma and leukemia. Although many translocations have been reported in the last four decades, the mechanism by which chromosomes break during a translocation remains largely unknown. In this review, we summarize recent advances made in understanding the molecular mechanism of chromosomal translocations.
42022861
Formation of a hyperdiploid karyotype in childhood acute lymphoblastic leukemia.
Hyperdiploidy with greater than or equal to 50 chromosomes is a frequent and distinct karyotypic pattern in the malignant cells of children with acute lymphoblastic leukemia. To understand better the mechanism of formation of the hyperdiploid karyotype, we studied 15 patients using 20 DNA probes that detect restriction fragment length polymorphisms. We first examined disomic chromosomes for loss of heterozygosity. Two patients had widespread loss of heterozygosity on all informative disomic chromosomes, and represent cases of near-haploid leukemia in which the chromosomes doubled. One other patient had loss of heterozygosity limited to chromosome 3; in this patient all of seven other informative disomic chromosomes retained heterozygosity. Loss of heterozygosity was not detected in the remaining 12 patients on a total of 87 informative disomic chromosomes. We then examined tetrasomic chromosomes for parental dosage. Of the 13 patients in whom widespread loss of heterozygosity was not present, 11 patients had tetrasomy 21; 10 of 11 (91%) had an equal dose of maternal and paternal alleles on chromosome 21 and only 1 of 11 (9%) had an unequal dose of parental alleles in a 3:1 ratio. These results suggest that the hyperdiploid karyotype usually arises by simultaneous gain of chromosomes from a diploid karyotype during a single abnormal cell division, and occasionally by doubling of chromosomes from a near-haploid karyotype. The hyperdiploidy in cases without widespread loss of heterozygosity is not caused by stepwise or sequential gains from a diploid karyotype or by losses from a tetraploid karyotype; the former should result in a 3:1 parental dosage for 67% of tetrasomic chromosomes (9% observed) and the latter should result in loss of heterozygosity for 33% of disomic chromosomes (1% observed). Additional studies of the molecular basis for this leukemia subtype are warranted.
28061417
Cell death by mitotic catastrophe: a molecular definition
The current literature is devoid of a clearcut definition of mitotic catastrophe, a type of cell death that occurs during mitosis. Here, we propose that mitotic catastrophe results from a combination of deficient cell-cycle checkpoints (in particular the DNA structure checkpoints and the spindle assembly checkpoint) and cellular damage. Failure to arrest the cell cycle before or at mitosis triggers an attempt of aberrant chromosome segregation, which culminates in the activation of the apoptotic default pathway and cellular demise. Cell death occurring during the metaphase/anaphase transition is characterized by the activation of caspase-2 (which can be activated in response to DNA damage) and/or mitochondrial membrane permeabilization with the release of cell death effectors such as apoptosis-inducing factor and the caspase-9 and-3 activator cytochrome c. Although the morphological aspect of apoptosis may be incomplete, these alterations constitute the biochemical hallmarks of apoptosis. Cells that fail to execute an apoptotic program in response to mitotic failure are likely to divide asymmetrically in the next round of cell division, with the consequent generation of aneuploid cells. This implies that disabling of the apoptotic program may actually favor chromosomal instability, through the suppression of mitotic catastrophe. Mitotic catastrophe thus may be conceived as a molecular device that prevents aneuploidization, which may participate in oncogenesis. Mitotic catastrophe is controlled by numerous molecular players, in particular, cell-cycle-specific kinases (such as the cyclin B1-dependent kinase Cdk1, polo-like kinases and Aurora kinases), cell-cycle checkpoint proteins, survivin, p53, caspases and members of the Bcl-2 family.
24295240
Programmed cell death in aging
Programmed cell death (PCD) pathways, including apoptosis and regulated necrosis, are required for normal cell turnover and tissue homeostasis. Mis-regulation of PCD is increasingly implicated in aging and aging-related disease. During aging the cell turnover rate declines for several highly-mitotic tissues. Aging-associated disruptions in systemic and inter-cell signaling combined with cell-autonomous damage and mitochondrial malfunction result in increased PCD in some cell types, and decreased PCD in other cell types. Increased PCD during aging is implicated in immune system decline, skeletal muscle wasting (sarcopenia), loss of cells in the heart, and neurodegenerative disease. In contrast, cancer cells and senescent cells are resistant to PCD, enabling them to increase in abundance during aging. PCD pathways limit life span in fungi, but whether PCD pathways normally limit adult metazoan life span is not yet clear. PCD is regulated by a balance of negative and positive factors, including the mitochondria, which are particularly subject to aging-associated malfunction.
5057
Ultrafast and memory-efficient alignment of short DNA sequences to the human genome
Bowtie is an ultrafast, memory-efficient alignment program for aligning short DNA sequence reads to large genomes. For the human genome, Burrows-Wheeler indexing allows Bowtie to align more than 25 million reads per CPU hour with a memory footprint of approximately 1.3 gigabytes. Bowtie extends previous Burrows-Wheeler techniques with a novel quality-aware backtracking algorithm that permits mismatches. Multiple processor cores can be used simultaneously to achieve even greater alignment speeds. Bowtie is open source http://bowtie.cbcb.umd.edu.
275837
COSMIC: exploring the world's knowledge of somatic mutations in human cancer
COSMIC, the Catalogue Of Somatic Mutations In Cancer (http://cancer.sanger.ac.uk) is the world's largest and most comprehensive resource for exploring the impact of somatic mutations in human cancer. Our latest release (v70; Aug 2014) describes 2 002 811 coding point mutations in over one million tumor samples and across most human genes. To emphasize depth of knowledge on known cancer genes, mutation information is curated manually from the scientific literature, allowing very precise definitions of disease types and patient details. Combination of almost 20 000 published studies gives substantial resolution of how mutations and phenotypes relate in human cancer, providing insights into the stratification of mutations and biomarkers across cancer patient populations. Conversely, our curation of cancer genomes (over 12 000) emphasizes knowledge breadth, driving discovery of unrecognized cancer-driving hotspots and molecular targets. Our high-resolution curation approach is globally unique, giving substantial insight into molecular biomarkers in human oncology. In addition, COSMIC also details more than six million noncoding mutations, 10 534 gene fusions, 61 299 genome rearrangements, 695 504 abnormal copy number segments and 60 119 787 abnormal expression variants. All these types of somatic mutation are annotated to both the human genome and each affected coding gene, then correlated across disease and mutation types.
8010036
Genome-wide analysis of non-coding regulatory mutations in cancer
Cancer primarily develops because of somatic alterations in the genome. Advances in sequencing have enabled large-scale sequencing studies across many tumor types, emphasizing the discovery of alterations in protein-coding genes. However, the protein-coding exome comprises less than 2% of the human genome. Here we analyze the complete genome sequences of 863 human tumors from The Cancer Genome Atlas and other sources to systematically identify noncoding regions that are recurrently mutated in cancer. We use new frequency- and sequence-based approaches to comprehensively scan the genome for noncoding mutations with potential regulatory impact. These methods identify recurrent mutations in regulatory elements upstream of PLEKHS1, WDR74 and SDHD, as well as previously identified mutations in the TERT promoter. SDHD promoter mutations are frequent in melanoma and are associated with reduced gene expression and poor prognosis. The non-protein-coding cancer genome remains widely unexplored, and our findings represent a step toward targeting the entire genome for clinical purposes.
21706427
Palimpsest: an R package for studying mutational and structural variant signatures along clonal evolution in cancer
Summary
Cancer genomes are altered by various mutational processes and, like palimpsests, bear the signatures of these different processes. The Palimpsest R package provides a complete workflow for the characterization and visualization of mutational signatures and their evolution along tumor development. The package covers a wide range of functions for extracting both base substitution and structural variant signatures, inferring the clonality of each alteration and analyzing the evolution of mutational processes between early clonal and late subclonal events. Palimpsest also estimates the probability of each mutation being due to each process to predict the mechanisms at the origin of driver events. Palimpsest is an easy-to-use toolset for reconstructing the natural history of a tumor using whole exome or whole genome sequencing data.


Availability and implementation
Palimpsest is freely available at www.github.com/FunGEST/Palimpsest.


Supplementary information
Supplementary data are available at Bioinformatics online.
7620813
Manta: rapid detection of structural variants and indels for germline and cancer sequencing applications
UNLABELLED
: We describe Manta, a method to discover structural variants and indels from next generation sequencing data. Manta is optimized for rapid germline and somatic analysis, calling structural variants, medium-sized indels and large insertions on standard compute hardware in less than a tenth of the time that comparable methods require to identify only subsets of these variant types: for example NA12878 at 50× genomic coverage is analyzed in less than 20 min. Manta can discover and score variants based on supporting paired and split-read evidence, with scoring models optimized for germline analysis of diploid individuals and somatic analysis of tumor-normal sample pairs. Call quality is similar to or better than comparable methods, as determined by pedigree consistency of germline calls and comparison of somatic calls to COSMIC database variants. Manta consistently assembles a higher fraction of its calls to base-pair resolution, allowing for improved downstream annotation and analysis of clinical significance. We provide Manta as a community resource to facilitate practical and routine structural variant analysis in clinical and research sequencing scenarios.


AVAILABILITY AND IMPLEMENTATION
Manta is released under the open-source GPLv3 license. Source code, documentation and Linux binaries are available from https://github.com/Illumina/manta.


CONTACT
csaunders@illumina.com


SUPPLEMENTARY INFORMATION
Supplementary data are available at Bioinformatics online.
4796595
Fast Computation and Applications of Genome Mappability
We present a fast mapping-based algorithm to compute the mappability of each region of a reference genome up to a specified number of mismatches. Knowing the mappability of a genome is crucial for the interpretation of massively parallel sequencing experiments. We investigate the properties of the mappability of eukaryotic DNA/RNA both as a whole and at the level of the gene family, providing for various organisms tracks which allow the mappability information to be visually explored. In addition, we show that mappability varies greatly between species and gene classes. Finally, we suggest several practical applications where mappability can be used to refine the analysis of high-throughput sequencing data (SNP calling, gene expression quantification and paired-end experiments). This work highlights mappability as an important concept which deserves to be taken into full account, in particular when massively parallel sequencing technologies are employed. The GEM mappability program belongs to the GEM (GEnome Multitool) suite of programs, which can be freely downloaded for any use from its website (http://gemlibrary.sourceforge.net).
20494277
ConsensusClusterPlus: a class discovery tool with confidence assessments and item tracking
Summary: Unsupervised class discovery is a highly useful technique in cancer research, where intrinsic groups sharing biological characteristics may exist but are unknown. The consensus clustering (CC) method provides quantitative and visual stability evidence for estimating the number of unsupervised classes in a dataset. ConsensusClusterPlus implements the CC method in R and extends it with new functionality and visualizations including item tracking, item-consensus and cluster-consensus plots. These new features provide users with detailed information that enable more specific decisions in unsupervised class discovery. Availability: ConsensusClusterPlus is open source software, written in R, under GPL-2, and available through the Bioconductor project (http://www.bioconductor.org/). Contact: mwilkers@med.unc.edu Supplementary Information: Supplementary data are available at Bioinformatics online.