DOCSLIB.ORG

Use of CRISPR-Modified Human Stem Cell Organoids to Study the Origin Of

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Use of CRISPR-Modified Human Stem Cell Organoids to Study the Origin Of RESEARCH CANCER MLH1 in normal human colonic organoids, we used CRISPR-Cas9 technology to insert a puromycin- resistance cassette into the second exon of the MLH1 gene (Fig. 1A and fig. S1A). After puromycin Use of CRISPR-modified human stem selection, we clonally expanded single organoids and subsequently genotyped these to confirm cor- cell organoids to study the origin of rect biallelic targeting (figs. S1, A and B, and S2A). Gene inactivation was verified by quantitative mutational signatures in cancer reverse transcription polymerase chain reaction (qRT-PCR), revealing a substantial reduction in MLH1 mRNA expression in MLH1 knockout Jarno Drost,1,2*† Ruben van Boxtel,2,3*† Francis Blokzijl,2,3 Tomohiro Mizutani,1,2 (MLH1KO) organoids, most likely due to the deg- Nobuo Sasaki,1,2 Valentina Sasselli,1,2 Joep de Ligt,2,3 Sam Behjati,4,5 radation (through nonsense-mediated decay) of Judith E. Grolleman,6 Tom van Wezel,7 Serena Nik-Zainal,4,8 Roland P. Kuiper,6,9 nonsense mRNAs transcribed from the mutant Edwin Cuppen,2,3‡ Hans Clevers1,2,9‡ alleles (Fig. 1C). Western blot analysis confirmed the loss of MLH1 protein expression in the MLH1KO Mutational processes underlie cancer initiation and progression. Signatures of these processes organoids (Fig. 1E). in cancer genomes may explain cancer etiology and could hold diagnostic and prognostic Next, we passaged the MLH1KO and parental value. We developed a strategy that can be used to explore the origin of cancer-associated normal human colon organoids for 2 months to mutational signatures. We used CRISPR-Cas9 technology to delete key DNA repair genes allow cells to accumulate sufficient mutations re- in human colon organoids, followed by delayed subcloning and whole-genome sequencing. We quired for downstream analyses (fig. S1C). We Downloaded from MLH1 found that mutation accumulation in organoids deficient in the mismatch repair gene then used flow cytometry to establish subclonal is driven by replication errors and accurately models the mutation profiles observed in cultures of single cells (fig. S1C) and expanded mismatch repair–deficient colorectal cancers. Application of this strategy to the cancer these until sufficient DNA could be obtained (7). NTHL1 predisposition gene , which encodes a base excision repair protein, revealed a Both clonal and subclonal cultures were subjected mutational footprint (signature 30) previously observed in a breast cancer cohort. We to whole-genome sequencing (WGS) analyses to NTHL1 show that signature 30 can arise from germline mutations. identify the mutations that accumulated between the two clonal expansion steps and to correlate http://science.sciencemag.org/ ancer arises through the sequential accu- repair (NER) pathways (5). However, multiple their appearance with time (Fig. 2A). As expected, mulation of mutations in somatic cells. The processes are simultaneously active in tumors MLH1KO organoids showed an increased base overallmutationalburdenofsomaticcells with highly unstable genomes, which makes it substitution load (27.7 ± 4.9 mutations per genome C is determined by a balance between DNA difficult to causally link the presence of specific per day) compared with normal organoids (3.8 ± damage and repair activity. Mutational mutational signatures to DNA repair deficiency. 1.2 mutations per genome per day), as well as a processes leave specific signatures, as defined Organoid technology allows for the long-term change in type of base substitutions (increase in by systematic analysis of mutation characteristics in vitro expansion of epithelial tissues, starting C>T transitions). Similarly, the MLH1KO organoids across many independent cancer sequencing data from a single adult stem cell. Such organoids re- displayed an increased number of INDELs (Fig. sets (1). A recent study showed that a set of mu- main genetically stable over long periods of time 2A), which were predominantly single–base-pair tational signatures in genome-wide mutation col- (6). We have previously shown that clonal organ- deletions (Fig. 2B) at mononucleotide repeats lections of breast cancers predicts deficiency of oid cultures can be used for in-depth pattern anal- (Fig. 2C). This confirmed that deletion of MLH1 on January 29, 2018 BRCA1 and BRCA2, as well as sensitivity to PARP ysis of mutations that accumulate throughout is sufficient to generate the mutator phenotype [poly(adenosine diphosphate–ribose) polymerase] life in tissue-specific adult stem cells (7, 8). More- observed in MMR-deficient tumors. As expected, inhibition (2). To date, the main approach to link over, organoids can be readily modified using we did not observe an increase in structural specific mutational signatures to underlying molec- CRISPR-Cas9 genome editing (9). Mammalian variations. ular mechanisms has involved associating cancer species differ greatly in their DNA repair capacity Somatic mutations are unevenly distributed mutations with defined exposures to carcinogens, and will thus respond differently to mutagenic throughout the genomes of most cancers (15, 16) such as tobacco smoke (3) and ionizing radiation stress (10, 11), and mutation types and numbers and normal adult stem cells (7). In general, base (4), or with the absence of specific DNA repair will vary over a lifetime (7). We used CRISPR- substitutions are more frequent in heterochro- proteins, such as components of the DNA mis- Cas9 genome editing in human intestinal organoid matic and late-replicating regions of the genome. match repair (MMR) (1) and nucleotide excision cultures to systematically decipher the mutational It has been reported that specific DNA repair ac- consequences of DNA repair deficiency. tivity, including MMR (17)andNER(18), under- 1Hubrecht Institute, Royal Netherlands Academy of Arts and To establish a tool for dissecting mutational lies this variation in regional mutation rates, as Sciences (KNAW) and University Medical Center (UMC) signatures in human organoid cultures, we in- tumors lacking essential components of either 2 Utrecht, 3584CT Utrecht, Netherlands. Cancer Genomics troduced loss-of-function mutations in the MMR of these DNA repair pathways show a less biased Netherlands, UMC Utrecht, 3584CX Utrecht, Netherlands. 3Center for Molecular Medicine, Department of Genetics, gene MutL homolog 1 (MLH1). Inactivating mu- genomic distribution. To directly test this, we per- UMC Utrecht, 3584CX Utrecht, Netherlands. 4Cancer tations in MMR genes, including MLH1,pre- formed a genome-wide analysis of replication Genome Project, Wellcome Trust Sanger Institute, Wellcome dispose people to colorectal cancer (12, 13). These timing (Repli-seq) (19) on human intestinal organ- Trust Genome Campus, Hinxton, Cambridgeshire CB10 1SA, tumors are characterized by an immense muta- oids. We defined early, intermediate, and late UK. 5Department of Paediatrics, University of Cambridge, Cambridge CB2 0QQ, UK. 6Department of Human Genetics, tional load, such as base substitutions and small replicating genomic regions and determined the Radboud Institute for Molecular Life Sciences, Radboud insertions and deletions (INDELs) at short repeat relationship between somatic mutational load University Medical Center, Nijmegen, Netherlands. sequences in the genome, referred to as micro- and replication timing. As observed in most can- 7 Departments of Pathology, Leiden University Medical satellite instability (MSI) (13). We used organoids cers (16), in vitro–accumulated mutations in Center, Leiden, Netherlands. 8East Anglian Medical Genetics Service, Cambridge University Hospitals National Health derived from human normal colon epithelium, normal organoids were more frequent in late- Service Foundation Trust, Cambridge, UK. 9Princess Máxima because this allowed us to study the effect of dis- replicating DNA (Fig. 3A). This bias was no longer Center for Pediatric Oncology, 3584CT Utrecht, Netherlands. ruption of single DNA repair genes in an otherwise present in MLH1KO cells, in line with the notion † *These authors contributed equally to this work. Present address: normal genetic background. Moreover, human thatMMRmightbemoreactiveineuchromatic Princess Máxima Center for Pediatric Oncology, 3584CT Utrecht, Netherlands. ‡Corresponding author. Email: ecuppen@ colonic organoids are as close as possible to the early-replicating regions of the genome, thereby umcutrecht.nl (E.C.); [email protected] (H.C.) cell-of-origin of colorectal cancer (14). To inactivate creating variation in regional mutation rates (17). Drost et al., Science 358, 234–238 (2017) 13 October 2017 1of5 RESEARCH | REPORT MLH1 locus exon 2 NTHL1 locus exon 2 sgRNA Targeting sgRNA strategy 5’ homology arm 3’ homology arm 5’ homology arm 3’ homology arm puromycin puromycin Homologous recombination puromycin puromycin MLH1 WT KO1 KO2 Downloaded from 1 1 MLH1 (~ 85kD) 70 * 55 tubulin (~55 kD) mRNA expression mRNA mRNA expression mRNA 0.5 0.5 NTHL1 http://science.sciencemag.org/ NTHL1 MLH1 WT KO4 KO5 KO6 35 NTHL1 (~34 kD) Relative Relative 0 0 wild-type MLH1 MLH1 wild-type NTHL1 NTHL1 NTHL1 knock-out 1 knock-out 2 knock-out 4 knock-out 5 knock-out 6 35 GAPDH (~37 kD) Fig. 1. Generation of DNA repair gene knock-outs in human intestinal independent experiments are indicated. (D)Sameasin(C),butforNTHL1. stem cell cultures. Targeting strategy for the generation of MLH1 (A)and (E) Western blot analysis of MLH1 expression in normal and MLH1KO organoids NTHL1 n (B) knockout organoids using CRISPR-Cas9 genome editing. sgRNA, (representative from = 3). Tubulin was
Load more

Recommended publications
  • Mutational Signatures: Emerging Concepts, Caveats and Clinical Applications
    REVIEWS Mutational signatures: emerging concepts, caveats and clinical applications Gene Koh 1,2, Andrea Degasperi 1,2, Xueqing Zou1,2, Sophie Momen1,2 and Serena Nik-Zainal 1,2 ✉ Abstract | Whole-genome sequencing has brought the cancer genomics community into new territory. Thanks to the sheer power provided by the thousands of mutations present in each patient’s cancer, we have been able to discern generic patterns of mutations, termed ‘mutational signatures’, that arise during tumorigenesis. These mutational signatures provide new insights into the causes of individual cancers, revealing both endogenous and exogenous factors that have influenced cancer development. This Review brings readers up to date in a field that is expanding in computational, experimental and clinical directions. We focus on recent conceptual advances, underscoring some of the caveats associated with using the mutational signature frameworks and highlighting the latest experimental insights. We conclude by bringing attention to areas that are likely to see advancements in clinical applications. The concept of mutational signatures was introduced or drug therapies, in an attempt to decipher causes7–9. in 2012 following the demonstration that analy­ However, the origins of many signatures remain cryp­ sis of all substitution mutations in a set of 21 whole­ tic. Furthermore, while earlier analyses reported single genome­sequenced (WGS) breast cancers could reveal signatures, for example of UV radiation (that is, SBS7)2, consistent patterns of mutagenesis across tumours1 more recent studies reported multiple versions of these (FIG. 1a). These patterns were the physiological imprints signatures (that is, SBS7a, SBS7b, SBS7c and SBS7d)3, of DNA damage and repair processes that had occurred leading the community to question whether some find­ during tumorigenesis and could distinguish BRCA1­null ings are reflective of biology or are simply mathemat­ and BRCA2­null tumours from sporadic breast cancers.
    [Show full text]
  • Trajectory and Uniqueness of Mutational Signatures in Yeast Mutators
    Trajectory and uniqueness of mutational signatures in yeast mutators Sophie Loeilleta,b, Mareike Herzogc, Fabio Pudduc, Patricia Legoixd, Sylvain Baulanded, Stephen P. Jacksonc, and Alain G. Nicolasa,b,1 aInstitut Curie, Paris Sciences et Lettres Research University, CNRS, UMR3244, 75248 Paris Cedex 05, France; bSorbonne Universités, Université Pierre et Marie Curie Paris 06, CNRS, UMR3244, 75248 Paris Cedex 05, France; cWellcome/Cancer Research UK Gurdon Institute and Department of Biochemistry, Cambridge CB2 1QN, United Kingdom; and dICGex NGS Platform, Institut Curie, 75248 Paris Cedex 05, France Edited by Richard D. Kolodner, Ludwig Institute for Cancer Research, La Jolla, CA, and approved August 24, 2020 (received for review June 2, 2020) The acquisition of mutations plays critical roles in adaptation, evolution, known (5, 15, 16). Here, we conducted a reciprocal functional ap- senescence, and tumorigenesis. Massive genome sequencing has proach to inactivate one or several genes involved in distinct ge- allowed extraction of specific features of many mutational landscapes nome maintenance processes (replication, repair, recombination, but it remains difficult to retrospectively determine the mechanistic oxidative stress response, or cell-cycle progression) in Saccharomy- origin(s), selective forces, and trajectories of transient or persistent ces cerevisiae diploids, establish the genome-wide mutational land- mutations and genome rearrangements. Here, we conducted a pro- scapes of mutation accumulation (MA) lines, explore the underlying spective reciprocal approach to inactivate 13 single or multiple evolu- mechanisms, and characterize the dynamics of mutation accumu- tionary conserved genes involved in distinct genome maintenance lation (and disappearance) along single-cell bottleneck passages. processes and characterize de novo mutations in 274 diploid Saccharo- myces cerevisiae mutation accumulation lines.
    [Show full text]
  • The Repertoire of Mutational Signatures in Human Cancer
    Article The repertoire of mutational signatures in human cancer https://doi.org/10.1038/s41586-020-1943-3 Ludmil B. Alexandrov1,25, Jaegil Kim2,25, Nicholas J. Haradhvala2,3,25, Mi Ni Huang4,5,25, Alvin Wei Tian Ng4,5, Yang Wu4,5, Arnoud Boot4,5, Kyle R. Covington6,7, Dmitry A. Gordenin8, Received: 18 May 2018 Erik N. Bergstrom1, S. M. Ashiqul Islam1, Nuria Lopez-Bigas9,10,11, Leszek J. Klimczak12, Accepted: 18 November 2019 John R. McPherson4,5, Sandro Morganella13, Radhakrishnan Sabarinathan10,14,15, David A. Wheeler6,16, Ville Mustonen17,18,19, PCAWG Mutational Signatures Working Group20, Published online: 5 February 2020 Gad Getz2,3,21,22,26, Steven G. Rozen4,5,23,26*, Michael R. Stratton13,26* & PCAWG Consortium24 The number of DBSs is proportional to the number of SBSs, with few exceptions Open access Somatic mutations in cancer genomes are caused by multiple mutational processes, each of which generates a characteristic mutational signature1. Here, as part of the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium2 of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA), we characterized mutational signatures using 84,729,690 somatic mutations from 4,645 whole-genome and 19,184 exome sequences that encompass most types of cancer. We identifed 49 single-base-substitution, 11 doublet-base-substitution, 4 clustered-base-substitution and 17 small insertion-and-deletion signatures. The substantial size of our dataset, compared with previous analyses3–15, enabled the discovery of new signatures, the separation of overlapping signatures and the decomposition of signatures into components that may represent associated—but distinct—DNA damage, repair and/or replication mechanisms.
    [Show full text]
  • Mutational Landscape in Genetically Engineered, Carcinogen- Induced, and Radiation-Induced Mouse Sarcoma
    Mutational landscape in genetically engineered, carcinogen- induced, and radiation-induced mouse sarcoma Chang-Lung Lee, … , Kouros Owzar, David G. Kirsch JCI Insight. 2019. https://doi.org/10.1172/jci.insight.128698. Research In-Press Preview Genetics Oncology Cancer development is influenced by hereditary mutations, somatic mutations due to random errors in DNA replication, or external factors. It remains unclear how distinct cell-intrinsic and -extrinsic factors impact oncogenesis within the same tissue type. We investigated murine soft tissue sarcomas generated by oncogenic alterations (KrasG12D activation and p53 deletion), carcinogens (3-methylcholanthrene [MCA] or ionizing radiation), and in a novel model combining both factors (MCA plus p53 deletion). Whole-exome sequencing demonstrated distinct mutational signatures in individual sarcoma cohorts. MCA-induced sarcomas exhibited high mutational burden and predominantly G-to-T transversions, while radiation-induced sarcomas exhibited low mutational burden and a distinct genetic signature characterized by C-to-T transitions. The indel to substitution ratio and amount of gene copy number variations were high for radiation-induced sarcomas. MCA-induced tumors generated on a p53-deficient background showed the highest genomic instability. MCA- induced sarcomas harbored mutations in putative cancer-driver genes that regulate MAPK signaling (Kras and Nf1) and the Hippo pathway (Fat1 and Fat4). In contrast, radiation-induced sarcomas and KrasG12Dp53–/– sarcomas did not harbor recurrent oncogenic mutations, rather they exhibited amplifications of specific oncogenes: Kras and Myc in KrasG12Dp53–/– sarcomas, and Met and Yap1 for radiation-induced sarcomas. These results reveal that different initiating events drive oncogenesis through distinct mechanisms. Find the latest version: https://jci.me/128698/pdf Mutational landscape in genetically engineered, carcinogen-induced, and radiation-induced mouse sarcoma Chang-Lung Lee1,2,3*, Yvonne M.
    [Show full text]
  • Clinical Outcome-Related Mutational Signatures Identified by Integrative
    Author Manuscript Published OnlineFirst on September 28, 2020; DOI: 10.1158/1078-0432.CCR-20-2854 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Clinical Outcome-Related Mutational Signatures Identified by Integrative Genomic Analysis in Nasopharyngeal Carcinoma Wei Dai1,2*, Dittman Lai-Shun Chung1, Larry Ka-Yue Chow1, Valen Zhuoyou Yu1, Lisa Chan Lei1, Merrin Man-Long Leong1, Candy King-Chi Chan1, Josephine Mun-Yee Ko1, Maria Li Lung1* 1Department of Clinical Oncology, University of Hong Kong, Hong Kong (SAR), P. R. China 2University of Hong Kong-Shenzhen Hospital, Shenzhen, P. R. China Running title: Clinical Outcome-Related Mutational Signatures in NPC *Co-corresponding authors: Maria Li Lung, Wei Dai MLL Address: Department of Clinical Oncology, The University of Hong Kong, Room L6- 43, 6/F, Laboratory Block, Faculty of Medicine Building, 21 Sassoon Road, Pokfulam, Hong Kong Email: [email protected] Tel: (852) 3917 9783 Fax: (852) 2816 6279 WD Address: Department of Clinical Oncology, The University of Hong Kong, Room L10- 56, 10/F, Laboratory Block, Faculty of Medicine Building, 21 Sassoon Road, Pokfulam, Hong Kong Email: [email protected] Tel: (852) 3917 6930 Fax: (852) 2816 6279 Conflict of interest The authors declare no potential conflicts of interest Translational relevance In NPC, general consensus for treatment is to use radiotherapy (RT) alone for stage I disease, RT with or without concurrent chemotherapy for stage II and chemoradiotherapy (CRT) for advanced stage disease. However, 15-58% of the cases do not respond well to conventional treatment, and, thus, have poor clinical outcomes.
    [Show full text]
  • Mutational Signatures: Experimental Design and Analytical Framework Gene Koh1,2,3†, Xueqing Zou2,3† and Serena Nik-Zainal2,3*
    Koh et al. Genome Biology (2020) 21:37 https://doi.org/10.1186/s13059-020-1951-5 REVIEW Open Access Mutational signatures: experimental design and analytical framework Gene Koh1,2,3†, Xueqing Zou2,3† and Serena Nik-Zainal2,3* Abstract Mutational signatures provide a powerful alternative for understanding the pathophysiology of cancer. Currently, experimental efforts aimed at validating and understanding the etiologies of cancer-derived mutational signatures are underway. In this review, we highlight key aspects of mutational signature experimental design and describe the analytical framework. We suggest guidelines and quality control measures for handling whole-genome sequencing data for mutational signature analyses and discuss pitfalls in interpretation. We envision that improved next-generation sequencing technologies and molecular cell biology approaches will usher in the next generation of studies into the etiologies and mechanisms of mutational patterns uncovered in cancers. Introduction In this review, we present guidelines that we hope will Somatic mutations arising through cell-intrinsic and ex- facilitate future experiments and analyses. We focus on ogenous processes mark the genome with distinctive pat- considerations in experimental design and on the com- terns termed mutational signatures. The field began in putational framework for data analysis in mutational sig- 2012 with the demonstration of at least 5 such mutation nature studies, particularly in human cellular model patterns in breast cancers [1]. Subsequently, 21 substitu- systems. We further discuss issues that need to be con- tion signatures were identifiable across 30 cancer types templated when linking an environmental mutagen or a [2]. While there have been revisions of analytical compo- DNA repair process to a mutational signature, which is nents of this field, there is a parallel trajectory evolving, fo- not as straightforward as may superficially seem.
    [Show full text]
  • Mutational Signature Analyses for Cancer Diagnostics Arne Van Hoeck1* , Niels H
    Hoeck et al. BMC Cancer (2019) 19:457 https://doi.org/10.1186/s12885-019-5677-2 REVIEW Open Access Portrait of a cancer: mutational signature analyses for cancer diagnostics Arne Van Hoeck1* , Niels H. Tjoonk1,2, Ruben van Boxtel2 and Edwin Cuppen1,3 Abstract Background: In the past decade, systematic and comprehensive analyses of cancer genomes have identified cancer driver genes and revealed unprecedented insight into the molecular mechanisms underlying the initiation and progression of cancer. These studies illustrate that although every cancer has a unique genetic make-up, there are only a limited number of mechanisms that shape the mutational landscapes of cancer genomes, as reflected by characteristic computationally-derived mutational signatures. Importantly, the molecular mechanisms underlying specific signatures can now be dissected and coupled to treatment strategies. Systematic characterization of mutational signatures in a cancer patient’s genome may thus be a promising new tool for molecular tumor diagnosis and classification. Results: In this review, we describe the status of mutational signature analysis in cancer genomes and discuss the opportunities and relevance, as well as future challenges, for further implementation of mutational signatures in clinical tumor diagnostics and therapy guidance. Conclusions: Scientific studies have illustrated the potential of mutational signature analysis in cancer research. As such, we believe that the implementation of mutational signature analysis within the diagnostic workflow will improve cancer diagnosis in the future. Keywords: Mutational signature, Cancer diagnosis, Cancer biomarkers, Cancer genomics, Molecular medicine, Whole genome sequencing Background promote tumorigenesis [2]. Genetic testing for driver Historically, cancer diagnostic and treatment decisions genes can identify the biological characteristics of tu- were predominantly based on tumor morphology, clin- mors.
    [Show full text]
  • Somatic Mutation Distributions in Cancer Genomes Vary with Three-Dimensional Chromatin Structure
    ARTICLES https://doi.org/10.1038/s41588-020-0708-0 Somatic mutation distributions in cancer genomes vary with three-dimensional chromatin structure Kadir C. Akdemir 1 ✉ , Victoria T. Le2, Justin M. Kim1,3, Sarah Killcoyne 4,5, Devin A. King6, Ya-Ping Lin7, Yanyan Tian8,9, Akira Inoue1, Samirkumar B. Amin 10, Frederick S. Robinson 11, Manjunath Nimmakayalu12, Rafael E. Herrera6, Erica J. Lynn8, Kin Chan13,14, Sahil Seth 11,15, Leszek J. Klimczak16, Moritz Gerstung 5, Dmitry A. Gordenin 13, John O’Brien 7, Lei Li8,17, Yonathan Lissanu Deribe1,18, Roel G. Verhaak 10, Peter J. Campbell19, Rebecca Fitzgerald 4, Ashby J. Morrison 6, Jesse R. Dixon 2 ✉ and P. Andrew Futreal 1 ✉ Somatic mutations in driver genes may ultimately lead to the development of cancer. Understanding how somatic mutations accumulate in cancer genomes and the underlying factors that generate somatic mutations is therefore crucial for develop- ing novel therapeutic strategies. To understand the interplay between spatial genome organization and specific mutational processes, we studied 3,000 tumor–normal-pair whole-genome datasets from 42 different human cancer types. Our analyses reveal that the change in somatic mutational load in cancer genomes is co-localized with topologically-associating-domain boundaries. Domain boundaries constitute a better proxy to track mutational load change than replication timing measure- ments. We show that different mutational processes lead to distinct somatic mutation distributions where certain processes generate mutations in active domains, and others generate mutations in inactive domains. Overall, the interplay between three-dimensional genome organization and active mutational processes has a substantial influence on the large-scale mutation-rate variations observed in human cancers.
    [Show full text]
  • Deficiency of Nucleotide Excision Repair Is Associated with Mutational Signature Observed in Cancer
    Downloaded from genome.cshlp.org on September 26, 2021 - Published by Cold Spring Harbor Laboratory Press Research Deficiency of nucleotide excision repair is associated with mutational signature observed in cancer Myrthe Jager,1,4 Francis Blokzijl,1,4,5 Ewart Kuijk,1 Johanna Bertl,2,6 Maria Vougioukalaki,3 Roel Janssen,1 Nicolle Besselink,1 Sander Boymans,1 Joep de Ligt,1 Jakob Skou Pedersen,2 Jan Hoeijmakers,3 Joris Pothof,3 Ruben van Boxtel,1,7 and Edwin Cuppen1 1Center for Molecular Medicine and Oncode Institute, University Medical Center Utrecht, Utrecht University, 3584 CG Utrecht, The Netherlands; 2Department of Molecular Medicine, Aarhus University, 8200 Aarhus N, Denmark; 3Erasmus Medical Center, 3015 CN Rotterdam, The Netherlands 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.
    [Show full text]
  • Mutational Signatures Are Markers of Drug Sensitivity of Cancer Cells
    bioRxiv preprint doi: https://doi.org/10.1101/2021.05.19.444811; this version posted May 21, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. Levatić et al. (2021) bioRxiv Mutational signatures are markers of drug sensitivity of cancer cells Jurica Levatić1, Marina Salvadores1, Francisco Fuster-Tormo1, Fran Supek1,2,* 1 Genome Data Science, Institute for Research in Biomedicine (IRB Barcelona), The Barcelona Institute of Science and Technology, C/ Baldiri Reixac 10, Barcelona 08028, Spain. 2 Catalan Institution for Advanced Studies (ICREA), Passeig de Lluís Companys, 23, 08010 Barcelona, Spain. * correspondence to: [email protected] Abstract Genomic analyses have revealed mutational signatures that are associated with DNA maintenance gone awry, a common occurrence in tumors. Because cancer therapeutics often target synthesis of DNA building blocks, DNA replication or DNA repair, we hypothesized that mutational signatures would make useful markers of drug sensitivity. We rigorously tested this hypothesis by a global analysis of various drug screening and genetic screening data sets, derived from cancer cell line panels. We introduce a novel computational method that detects mutational signatures in cell lines by stringently adjusting for the confounding germline mutational processes, which are difficult to remove when healthy samples from the same individuals are not available. This revealed many associations between diverse mutational signatures and drug activity in cancer cell lines, which are comparably or more numerous than associations with classical genetic features such as cancer driver mutations or copy number alterations.
    [Show full text]
  • Learning Mutational Signatures and Their Multidimensional Genomic
    ARTICLE https://doi.org/10.1038/s41467-021-23551-9 OPEN Learning mutational signatures and their multidimensional genomic properties with TensorSignatures ✉ Harald Vöhringer1, Arne Van Hoeck 2, Edwin Cuppen2,3 & Moritz Gerstung 1,4 We present TensorSignatures, an algorithm to learn mutational signatures jointly across different variant categories and their genomic localisation and properties. The analysis of 1234567890():,; 2778 primary and 3824 metastatic cancer genomes of the PCAWG consortium and the HMF cohort shows that all signatures operate dynamically in response to genomic states. The analysis pins differential spectra of UV mutagenesis found in active and inactive chromatin to global genome nucleotide excision repair. TensorSignatures accurately characterises transcription-associated mutagenesis in 7 different cancer types. The algorithm also extracts distinct signatures of replication- and double strand break repair-driven mutagenesis by APOBEC3A and 3B with differential numbers and length of mutation clusters. Finally, Ten- sorSignatures reproduces a signature of somatic hypermutation generating highly clustered variants at transcription start sites of active genes in lymphoid leukaemia, distinct from a general and less clustered signature of Polη-driven translesion synthesis found in a broad range of cancer types. In summary, TensorSignatures elucidates complex mutational foot- prints by characterising their underlying processes with respect to a multitude of genomic variables. 1 European Molecular Biology Laboratory, European
    [Show full text]
  • 1 Mutational Signatures of DNA Mismatch Repair Deficiency in C
    bioRxiv preprint doi: https://doi.org/10.1101/149153; this version posted June 13, 2017. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Mutational signatures of DNA mismatch repair deficiency in C. elegans and human cancers Authors: Meier B*1, Volkova N*2, Hong Y1, Schofield P1,3, Campbell PJ 4, 5, 6 Gerstung M2 and A Gartner1 * these authors contributed equally. Institutions: 1 Centre for Gene Regulation and Expression, University of Dundee, Dundee, UK. 2 European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL- EBI), Hinxton, Cambridgeshire, UK. 3 Division of Computational Biology, University of Dundee, Dundee, UK. 4 Cancer Genome Project, Wellcome Trust Sanger Institute, Hinxton, Cambridge, UK. 5 Department of Haematology, University of Cambridge, Cambridge, UK. 6 Department of Haematology, Addenbrooke’s Hospital, Cambridge, UK. Correspondence: Dr Anton Gartner Centre for Gene Regulation and Expression University of Dundee Dow Street Dundee, DD1 5EH UK. Tel: +44 (0) 1382 385809 E-mail: [email protected] Dr Moritz Gerstung European Molecular Biology Laboratory European Bioinformatics Institute (EMBL-EBI) Hinxton, CB10 1SA Cambridgeshire UK. Tel: +44 (0) 1223 494636 E-mail: [email protected] Keywords: DNA mismatch repair, mutational signatures, hypermutation, replication errors, POLE-4, genome integrity, C. elegans 1 bioRxiv preprint doi: https://doi.org/10.1101/149153; this version posted June 13, 2017. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
    [Show full text]