Publications by authors named "Edwin Cuppen"

246 Publications

Limited evolution of the actionable metastatic cancer genome under therapeutic pressure.

Nat Med 2021 09 9;27(9):1553-1563. Epub 2021 Aug 9.

Division of Molecular Oncology & Immunology, Netherlands Cancer Institute, Amsterdam, the Netherlands.

Genomic profiling is critical for the identification of treatment options for patients with metastatic cancer, but it remains unclear how frequently this procedure should be repeated during the course of the disease. To address this, we analyzed whole-genome sequencing (WGS) data of 250 biopsy pairs, longitudinally collected over the treatment course of 231 adult patients with a representative variety of metastatic solid malignancies. Within the biopsy interval (median, 6.4 months), patients received one or multiple lines of (mostly) standard-of-care (SOC) treatments, with all major treatment modalities being broadly represented. SOC biomarkers and biomarkers for clinical trial enrollment could be identified in 23% and 72% of biopsies, respectively. For SOC genomic biomarkers, we observed full concordance between the first and the second biopsy in 99% of pairs. Of the 219 biomarkers for clinical trial enrollment that were identified in the first biopsies, we recovered 94% in the follow-up biopsies. Furthermore, a second WGS analysis did not identify additional biomarkers for clinical trial enrollment in 91% of patients. More-frequent genomic evolution was observed when considering specific genes targeted by small-molecule inhibitors or hormonal therapies (21% and 22% of cases, respectively). Together, our data demonstrate that there is limited evolution of the actionable genome of treated metastases. A single WGS analysis of a metastatic biopsy is generally sufficient to identify SOC genomic biomarkers and to identify investigational treatment opportunities.
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http://dx.doi.org/10.1038/s41591-021-01448-wDOI Listing
September 2021

The genomic landscape of 85 advanced neuroendocrine neoplasms reveals subtype-heterogeneity and potential therapeutic targets.

Nat Commun 2021 07 29;12(1):4612. Epub 2021 Jul 29.

Department of Medical Oncology, Erasmus MC Cancer Institute, Rotterdam, the Netherlands.

Metastatic and locally-advanced neuroendocrine neoplasms (aNEN) form clinically and genetically heterogeneous malignancies, characterized by distinct prognoses based upon primary tumor localization, functionality, grade, proliferation index and diverse outcomes to treatment. Here, we report the mutational landscape of 85 whole-genome sequenced aNEN. This landscape reveals distinct genomic subpopulations of aNEN based on primary localization and differentiation grade; we observe relatively high tumor mutational burdens (TMB) in neuroendocrine carcinoma (average 5.45 somatic mutations per megabase) with TP53, KRAS, RB1, CSMD3, APC, CSMD1, LRATD2, TRRAP and MYC as major drivers versus an overall low TMB in neuroendocrine tumors (1.09). Furthermore, we observe distinct drivers which are enriched in somatic aberrations in pancreatic (MEN1, ATRX, DAXX, DMD and CREBBP) and midgut-derived neuroendocrine tumors (CDKN1B). Finally, 49% of aNEN patients reveal potential therapeutic targets based upon actionable (and responsive) somatic aberrations within their genome; potentially directing improvements in aNEN treatment strategies.
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http://dx.doi.org/10.1038/s41467-021-24812-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8322054PMC
July 2021

Clinical interpretation of whole-genome and whole-transcriptome sequencing for precision oncology.

Semin Cancer Biol 2021 Jul 10. Epub 2021 Jul 10.

Institute for Computational Biomedicine, Weill Cornell Medicine, New York, United States; Englander Institute for Precision Medicine, Weill Cornell Medicine, New York, United States. Electronic address:

Whole-genome sequencing either alone or in combination with whole-transcriptome sequencing has started to be used to analyze clinical tumor samples to improve diagnosis, provide risk stratification, and select patient-specific therapies. Compared with current genomic testing strategies, largely focused on small number of genes tested individually or targeted panels, whole-genome and transcriptome sequencing (WGTS) provides novel opportunities to identify and report a potentially much larger number of actionable alterations with diagnostic, prognostic, and/or predictive impact. Such alterations include point mutations, indels, copy- number aberrations and structural variants, but also germline variants, fusion genes, noncoding alterations and mutational signatures. Nevertheless, these comprehensive tests are accompanied by many challenges ranging from the extent and diversity of sequence alterations detected by these methods to the complexity and limited existing standardization in interpreting them. We describe the challenges of WGTS interpretation and the opportunities with comprehensive genomic testing.
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http://dx.doi.org/10.1016/j.semcancer.2021.07.003DOI Listing
July 2021

GRIDSS2: comprehensive characterisation of somatic structural variation using single breakend variants and structural variant phasing.

Genome Biol 2021 07 12;22(1):202. Epub 2021 Jul 12.

Bioinformatics Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Australia.

GRIDSS2 is the first structural variant caller to explicitly report single breakends-breakpoints in which only one side can be unambiguously determined. By treating single breakends as a fundamental genomic rearrangement signal on par with breakpoints, GRIDSS2 can explain 47% of somatic centromere copy number changes using single breakends to non-centromere sequence. On a cohort of 3782 deeply sequenced metastatic cancers, GRIDSS2 achieves an unprecedented 3.1% false negative rate and 3.3% false discovery rate and identifies a novel 32-100 bp duplication signature. GRIDSS2 simplifies complex rearrangement interpretation through phasing of structural variants with 16% of somatic calls phasable using paired-end sequencing.
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http://dx.doi.org/10.1186/s13059-021-02423-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8274009PMC
July 2021

Reconstructing single-cell karyotype alterations in colorectal cancer identifies punctuated and gradual diversification patterns.

Nat Genet 2021 08 1;53(8):1187-1195. Epub 2021 Jul 1.

Molecular Cancer Research, Center for Molecular Medicine, University Medical Center Utrecht, Utrecht University, Utrecht, the Netherlands.

Central to tumor evolution is the generation of genetic diversity. However, the extent and patterns by which de novo karyotype alterations emerge and propagate within human tumors are not well understood, especially at single-cell resolution. Here, we present 3D Live-Seq-a protocol that integrates live-cell imaging of tumor organoid outgrowth and whole-genome sequencing of each imaged cell to reconstruct evolving tumor cell karyotypes across consecutive cell generations. Using patient-derived colorectal cancer organoids and fresh tumor biopsies, we demonstrate that karyotype alterations of varying complexity are prevalent and can arise within a few cell generations. Sub-chromosomal acentric fragments were prone to replication and collective missegregation across consecutive cell divisions. In contrast, gross genome-wide karyotype alterations were generated in a single erroneous cell division, providing support that aneuploid tumor genomes can evolve via punctuated evolution. Mapping the temporal dynamics and patterns of karyotype diversification in cancer enables reconstructions of evolutionary paths to malignant fitness.
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http://dx.doi.org/10.1038/s41588-021-00891-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8346364PMC
August 2021

Clinical utility of whole-genome sequencing in precision oncology.

Semin Cancer Biol 2021 Jun 25. Epub 2021 Jun 25.

NIHR Oxford Biomedical Research Centre and Department of Oncology, University of Oxford, Oxford, United Kingdom. Electronic address:

Precision diagnostics is one of the two pillars of precision medicine. Sequencing efforts in the past decade have firmly established cancer as a primarily genetically driven disease. This concept is supported by therapeutic successes aimed at particular pathways that are perturbed by specific driver mutations in protein-coding domains and reflected in three recent FDA tissue agnostic cancer drug approvals. In addition, there is increasing evidence from studies that interrogate the entire genome by whole-genome sequencing that acquired global and complex genomic aberrations including those in non-coding regions of the genome might also reflect clinical outcome. After addressing technical, logistical, financial and ethical challenges, national initiatives now aim to introduce clinical whole-genome sequencing into real-world diagnostics as a rational and potentially cost-effective tool for response prediction in cancer and to identify patients who would benefit most from 'expensive' targeted therapies and recruitment into clinical trials. However, so far, this has not been accompanied by a systematic and prospective evaluation of the clinical utility of whole-genome sequencing within clinical trials of uniformly treated patients of defined clinical outcome. This approach would also greatly facilitate novel predictive biomarker discovery and validation, ultimately reducing size and duration of clinical trials and cost of drug development. This manuscript is the third in a series of three to review and critically appraise the potential and challenges of clinical whole-genome sequencing in solid tumors and hematological malignancies.
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http://dx.doi.org/10.1016/j.semcancer.2021.06.018DOI Listing
June 2021

Learning mutational signatures and their multidimensional genomic properties with TensorSignatures.

Nat Commun 2021 06 15;12(1):3628. Epub 2021 Jun 15.

European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL-EBI), Hinxton, UK.

We present TensorSignatures, an algorithm to learn mutational signatures jointly across different variant categories and their genomic localisation and properties. The analysis of 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, TensorSignatures 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 footprints by characterising their underlying processes with respect to a multitude of genomic variables.
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http://dx.doi.org/10.1038/s41467-021-23551-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8206343PMC
June 2021

Analytical demands to use whole-genome sequencing in precision oncology.

Semin Cancer Biol 2021 Jun 10. Epub 2021 Jun 10.

MLL Munich Leukemia Laboratory, Munich, Germany. Electronic address:

Interrogating the tumor genome in its entirety by whole-genome sequencing (WGS) offers an unprecedented insight into the biology and pathogenesis of cancer, with potential impact on diagnostics, prognostication and therapy selection. WGS is able to detect sequence as well as structural variants and thereby combines central domains of cytogenetics and molecular genetics. Given the potential of WGS in directing targeted therapeutics and clinical decision-making, we envision a gradual transition of the method from research to clinical routine. This review is one out of three within this issue aimed at facilitating this effort, by discussing in-depth analytical validation, clinical interpretation and clinical utility of WGS. The review highlights the requirements for implementing, validating and maintaining a clinical WGS pipeline to obtain high-quality patient-specific data in accordance with the local regulatory landscape. Every step of the WGS pipeline, which includes DNA extraction, library preparation, sequencing, bioinformatics analysis, and data storage, is considered with respect to its logistics, necessities, potential pitfalls, and the required quality management. WGS is likely to drive clinical diagnostics and patient care forward, if requirements and challenges of the technique are recognized and met.
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http://dx.doi.org/10.1016/j.semcancer.2021.06.009DOI Listing
June 2021

VIRUSBreakend: Viral Integration Recognition Using Single Breakends.

Bioinformatics 2021 May 11. Epub 2021 May 11.

Bioinformatics Division, Walter and Eliza Hall Institute of Medical Research, Parkville, Australia.

Motivation: Integration of viruses into infected host cell DNA can cause DNA damage and disrupt genes. Recent cost reductions and growth of whole genome sequencing has produced a wealth of data in which viral presence and integration detection is possible. While key research and clinically relevant insights can be uncovered, existing software has not achieved widespread adoption, limited in part due to high computational costs, the inability to detect a wide range of viruses, as well as precision and sensitivity.

Results: Here, we describe VIRUSBreakend, a high-speed tool that identifies viral DNA presence and genomic integration. It utilizes single breakends, breakpoints in which only one side can be unambiguously placed, in a novel virus-centric variant calling and assembly approach to identify viral integrations with high sensitivity and a near-zero false discovery rate. VIRUSBreakend detects viral integrations anywhere in the host genome including regions such as centromeres and telomeres unable to be called by existing tools. Applying VIRUSBreakend to a large metastatic cancer cohort, we demonstrate that it can reliably detect clinically relevant viral presence and integration including HPV, HBV, MCPyV, EBV, and HHV-8.

Availability: VIRUSBreakend is part of the Genomic Rearrangement IDentification Software Suite (GRIDSS). It is available under a GPLv3 license from https://github.com/PapenfussLab/VIRUSBreakend.

Supplementary Information: Supplementary data are available at Bioinformatics online.
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http://dx.doi.org/10.1093/bioinformatics/btab343DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8504616PMC
May 2021

Clinical Validation of Whole Genome Sequencing for Cancer Diagnostics.

J Mol Diagn 2021 07 6;23(7):816-833. Epub 2021 May 6.

Hartwig Medical Foundation, Amsterdam, the Netherlands; Center for Molecular Medicine and Oncode Institute, University Medical Center Utrecht, Utrecht, the Netherlands.

Whole genome sequencing (WGS) using fresh-frozen tissue and matched blood samples from cancer patients may become the most complete genetic tumor test. With the increasing availability of small biopsies and the need to screen more number of biomarkers, the use of a single all-inclusive test is preferable over multiple consecutive assays. To meet high-quality diagnostics standards, we optimized and clinically validated WGS sample and data processing procedures, resulting in a technical success rate of 95.6% for fresh-frozen samples with sufficient (≥20%) tumor content. Independent validation of identified biomarkers against commonly used diagnostic assays showed a high sensitivity (recall; 98.5%) and precision (positive predictive value; 97.8%) for detection of somatic single-nucleotide variants and insertions and deletions (across 22 genes), and high concordance for detection of gene amplification (97.0%; EGFR and MET) as well as somatic complete loss (100%; CDKN2A/p16). Gene fusion analysis showed a concordance of 91.3% between DNA-based WGS and an orthogonal RNA-based gene fusion assay. Microsatellite (in)stability assessment showed a sensitivity of 100% with a precision of 94%, and virus detection (human papillomavirus), an accuracy of 100% compared with standard testing. In conclusion, whole genome sequencing has a >95% sensitivity and precision compared with routinely used DNA techniques in diagnostics, and all relevant mutation types can be detected reliably in a single assay.
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http://dx.doi.org/10.1016/j.jmoldx.2021.04.011DOI Listing
July 2021

Early technology assessment of using whole genome sequencing in personalized oncology.

Expert Rev Pharmacoecon Outcomes Res 2021 Jun 1;21(3):343-351. Epub 2021 Jun 1.

Health Technology and Services Research Department, Technical Medical Centre, University of Twente, Enschede, The Netherlands.

: Personalized medicine-based treatments in advanced cancer hold the promise to offer substantial health benefits to genetic subgroups, but require efficient biomarker-based patient stratification to match the right treatment and may be expensive. Standard molecular diagnostics are currently very heterogeneous, and tests are often performed sequentially. The alternative to whole genome sequencing (WGS) i.e. simultaneously testing for all relevant DNA-based biomarkers thereby allowing immediate selection of the most optimal therapy, is more costly than current techniques. In the current implementation stage, it is important to explore the added value and cost-effectiveness of using WGS on a patient level and to assess optimal introduction of WGS on the level of the healthcare system.: First, an overview of current worldwide initiatives concerning the use of WGS in clinical practice for cancer diagnostics is given. Second, a comprehensive, early health technology assessment (HTA) approach of evaluating WGS in the Netherlands is described, relating to the following aspects: diagnostic value, WGS-based treatment decisions, assessment of long-term health benefits and harms, early cost-effectiveness modeling, nation-wide organization, and Ethical, Legal and Societal Implications.: This study provides evidence to guide further development and implementation of WGS in clinical practice and the healthcare system.
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http://dx.doi.org/10.1080/14737167.2021.1917386DOI Listing
June 2021

Micro-costing diagnostics in oncology: from single-gene testing to whole- genome sequencing.

Expert Rev Pharmacoecon Outcomes Res 2021 Jun 6;21(3):413-414. Epub 2021 May 6.

Julius Center for Health Sciences and Primary Care, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands.

: Predictive diagnostics play an increasingly important role in personalized medicine for cancer treatment. Whole-genome sequencing (WGS)-based treatment selection is expected to rapidly increase worldwide. This study aimed to calculate and compare the total cost of currently used diagnostic techniques and of WGS in treatment of non-small cell lung carcinoma (NSCLC), melanoma, colorectal cancer (CRC), and gastrointestinal stromal tumor (GIST) in the Netherlands.: The activity-based costing (ABC) method was conducted to calculate total cost of included diagnostic techniques based on data provided by Dutch pathology laboratories and the Dutch-centralized cancer WGS facility. Costs were allocated to four categories: capital costs, maintenance costs, software costs, and operational costs.: The total cost per cancer patient per technique varied from € 58 (Sanger sequencing, three amplicons) to € 2925 (paired tumor-normal WGS). The operational costs accounted for the vast majority (over 90%) of the total per cancer patient technique costs.: This study outlined in detail all costing aspects and cost prices of current and new diagnostic modalities used in treatment of NSCLC, melanoma, CRC, and GIST in the Netherlands. Detailed cost differences and value comparisons between these diagnostic techniques enable future economic evaluations to support decision-making.
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http://dx.doi.org/10.1080/14737167.2021.1917385DOI Listing
June 2021

Whole genome sequencing of metastatic colorectal cancer reveals prior treatment effects and specific metastasis features.

Nat Commun 2021 01 25;12(1):574. Epub 2021 Jan 25.

Department of Medical Oncology, Erasmus MC Cancer Institute, Erasmus University Medical Center Rotterdam, Rotterdam, The Netherlands.

In contrast to primary colorectal cancer (CRC) little is known about the genomic landscape of metastasized CRC. Here we present whole genome sequencing data of metastases of 429 CRC patients participating in the pan-cancer CPCT-02 study (NCT01855477). Unsupervised clustering using mutational signature patterns highlights three major patient groups characterized by signatures known from primary CRC, signatures associated with received prior treatments, and metastasis-specific signatures. Compared to primary CRC, we identify additional putative (non-coding) driver genes and increased frequencies in driver gene mutations. In addition, we identify specific genes preferentially affected by microsatellite instability. CRC-specific 1kb-10Mb deletions, enriched for common fragile sites, and LINC00672 mutations are associated with response to treatment in general, whereas FBXW7 mutations predict poor response specifically to EGFR-targeted treatment. In conclusion, the genomic landscape of mCRC shows defined changes compared to primary CRC, is affected by prior treatments and contains features with potential clinical relevance.
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http://dx.doi.org/10.1038/s41467-020-20887-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7835235PMC
January 2021

Human extrahepatic and intrahepatic cholangiocyte organoids show region-specific differentiation potential and model cystic fibrosis-related bile duct disease.

Sci Rep 2020 12 14;10(1):21900. Epub 2020 Dec 14.

Department of Surgery, Erasmus MC-University Medical Center, Wytemaweg 80, 3015 CN, Rotterdam, The Netherlands.

The development, homeostasis, and repair of intrahepatic and extrahepatic bile ducts are thought to involve distinct mechanisms including proliferation and maturation of cholangiocyte and progenitor cells. This study aimed to characterize human extrahepatic cholangiocyte organoids (ECO) using canonical Wnt-stimulated culture medium previously developed for intrahepatic cholangiocyte organoids (ICO). Paired ECO and ICO were derived from common bile duct and liver tissue, respectively. Characterization showed both organoid types were highly similar, though some differences in size and gene expression were observed. Both ECO and ICO have cholangiocyte fate differentiation capacity. However, unlike ICO, ECO lack the potential for differentiation towards a hepatocyte-like fate. Importantly, ECO derived from a cystic fibrosis patient showed no CFTR channel activity but normal chloride channel and MDR1 transporter activity. In conclusion, this study shows that ECO and ICO have distinct lineage fate and that ECO provide a competent model to study extrahepatic bile duct diseases like cystic fibrosis.
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http://dx.doi.org/10.1038/s41598-020-79082-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7736890PMC
December 2020

Study protocol: Whole genome sequencing Implementation in standard Diagnostics for Every cancer patient (WIDE).

BMC Med Genomics 2020 11 10;13(1):169. Epub 2020 Nov 10.

Department of Pathology, Netherlands Cancer Institute, Amsterdam, The Netherlands.

Background: 'Precision oncology' can ensure the best suitable treatment at the right time by tailoring treatment towards individual patient and comprehensive tumour characteristics. In current molecular pathology, diagnostic tests which are part of the standard of care (SOC) only cover a limited part of the spectrum of genomic changes, and often are performed in an iterative way. This occurs at the expense of valuable patient time, available tissue sample, and interferes with 'first time right' treatment decisions. Whole Genome Sequencing (WGS) captures a near complete view of genomic characteristics of a tumour in a single test. Moreover, WGS facilitates faster implementation of new treatment relevant biomarkers. At present, WGS mainly has been applied in study settings, but its performance in a routine diagnostic setting remains to be evaluated. The WIDE study aims to investigate the feasibility and validity of WGS-based diagnostics in clinical practice.

Methods: 1200 consecutive patients in a single comprehensive cancer centre with (suspicion of) a metastasized solid tumour will be enrolled with the intention to analyse tumour tissue with WGS, in parallel to SOC diagnostics. Primary endpoints are (1) feasibility of implementation of WGS-based diagnostics into routine clinical care and (2) clinical validation of WGS by comparing identification of treatment-relevant variants between WGS and SOC molecular diagnostics. Secondary endpoints entail (1) added clinical value in terms of additional treatment options and (2) cost-effectiveness of WGS compared to SOC diagnostics through a Health Technology Assessment (HTA) analysis. Furthermore, the (3) perceived impact of WGS-based diagnostics on clinical decision making will be evaluated through questionnaires. The number of patients included in (experimental) therapies initiated based on SOC or WGS diagnostics will be reported with at least 3 months follow-up. The clinical efficacy is beyond the scope of WIDE. Key performance indicators will be evaluated after every 200 patients enrolled, and procedures optimized accordingly, to continuously improve the diagnostic performance of WGS in a routine clinical setting.

Discussion: WIDE will yield the optimal conditions under which WGS can be implemented in a routine molecular diagnostics setting and establish the position of WGS compared to SOC diagnostics in routine clinical care.
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http://dx.doi.org/10.1186/s12920-020-00814-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7654005PMC
November 2020

Pan-cancer landscape of homologous recombination deficiency.

Nat Commun 2020 11 4;11(1):5584. Epub 2020 Nov 4.

Center for Molecular Medicine and Oncode Institute, University Medical Center Utrecht, Utrecht, The Netherlands.

Homologous recombination deficiency (HRD) results in impaired double strand break repair and is a frequent driver of tumorigenesis. Here, we develop a genome-wide mutational scar-based pan-cancer Classifier of HOmologous Recombination Deficiency (CHORD) that can discriminate BRCA1- and BRCA2-subtypes. Analysis of a metastatic (n = 3,504) and primary (n = 1,854) pan-cancer cohort reveals that HRD is most frequent in ovarian and breast cancer, followed by pancreatic and prostate cancer. We identify biallelic inactivation of BRCA1, BRCA2, RAD51C or PALB2 as the most common genetic cause of HRD, with RAD51C and PALB2 inactivation resulting in BRCA2-type HRD. We find that while the specific genetic cause of HRD is cancer type specific, biallelic inactivation is predominantly associated with loss-of-heterozygosity (LOH), with increased contribution of deep deletions in prostate cancer. Our results demonstrate the value of pan-cancer genomics-based HRD testing and its potential diagnostic value for patient stratification towards treatment with e.g. poly ADP-ribose polymerase inhibitors (PARPi).
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http://dx.doi.org/10.1038/s41467-020-19406-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7643118PMC
November 2020

Single-cell derived tumor organoids display diversity in HLA class I peptide presentation.

Nat Commun 2020 10 21;11(1):5338. Epub 2020 Oct 21.

Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands.

Tumor heterogeneity is a major cause of therapeutic resistance. Immunotherapy may exploit alternative vulnerabilities of drug-resistant cells, where tumor-specific human leukocyte antigen (HLA) peptide ligands are promising leads to invoke targeted anti-tumor responses. Here, we investigate the variability in HLA class I peptide presentation between different clonal cells of the same colorectal cancer patient, using an organoid system. While clone-specific differences in HLA peptide presentation were observed, broad inter-clone variability was even more prevalent (15-25%). By coupling organoid proteomics and HLA peptide ligandomics, we also found that tumor-specific ligands from DNA damage control and tumor suppressor source proteins were prominently presented by tumor cells, coinciding likely with the silencing of such cytoprotective functions. Collectively, these data illustrate the heterogeneous HLA peptide presentation landscape even within one individual, and hint that a multi-peptide vaccination approach against highly conserved tumor suppressors may be a viable option in patients with low tumor-mutational burden.
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http://dx.doi.org/10.1038/s41467-020-19142-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7577990PMC
October 2020

Publisher Correction: The mutational impact of culturing human pluripotent and adult stem cells.

Nat Commun 2020 08 4;11(1):3932. Epub 2020 Aug 4.

Center for Molecular Medicine and Oncode Institute, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG, Utrecht, The Netherlands.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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http://dx.doi.org/10.1038/s41467-020-17797-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7403145PMC
August 2020

Patient-Derived Ovarian Cancer Organoids Mimic Clinical Response and Exhibit Heterogeneous Inter- and Intrapatient Drug Responses.

Cell Rep 2020 06;31(11):107762

Genetics, Center for Molecular Medicine, University Medical Center Utrecht, Utrecht University, Universiteitsweg 100, 3584 CG Utrecht, the Netherlands; Oncode Institute, Jaarbeursplein 6, 3521 AL Utrecht, the Netherlands. Electronic address:

There remains an unmet need for preclinical models to enable personalized therapy for ovarian cancer (OC) patients. Here we evaluate the capacity of patient-derived organoids (PDOs) to predict clinical drug response and functional consequences of tumor heterogeneity. We included 36 whole-genome-characterized PDOs from 23 OC patients with known clinical histories. OC PDOs maintain the genomic features of the original tumor lesion and recapitulate patient response to neoadjuvant carboplatin/paclitaxel combination treatment. PDOs display inter- and intrapatient drug response heterogeneity to chemotherapy and targeted drugs, which can be partially explained by genetic aberrations. PDO drug screening identifies high responsiveness to at least one drug for 88% of patients. PDOs are valuable preclinical models that can provide insights into drug response for individual patients with OC, complementary to genetic testing. Generating PDOs of multiple tumor locations can improve clinical decision making and increase our knowledge of genetic and drug response heterogeneity.
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http://dx.doi.org/10.1016/j.celrep.2020.107762DOI Listing
June 2020

Sperm DNA damage causes genomic instability in early embryonic development.

Sci Adv 2020 Apr 15;6(16):eaaz7602. Epub 2020 Apr 15.

Center for Molecular Medicine and Oncode Institute, University Medical Center Utrecht, Utrecht University, Universiteitsweg 100, Utrecht 3584 CG, Netherlands.

Genomic instability is common in human embryos, but the underlying causes are largely unknown. Here, we examined the consequences of sperm DNA damage on the embryonic genome by single-cell whole-genome sequencing of individual blastomeres from bovine embryos produced with sperm damaged by γ-radiation. Sperm DNA damage primarily leads to fragmentation of the paternal chromosomes followed by random distribution of the chromosomal fragments over the two sister cells in the first cell division. An unexpected secondary effect of sperm DNA damage is the induction of direct unequal cleavages, which include the poorly understood heterogoneic cell divisions. As a result, chaotic mosaicism is common in embryos derived from fertilizations with damaged sperm. The mosaic aneuploidies, uniparental disomies, and de novo structural variation induced by sperm DNA damage may compromise fertility and lead to rare congenital disorders when embryos escape developmental arrest.
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http://dx.doi.org/10.1126/sciadv.aaz7602DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7159919PMC
April 2020

The mutational impact of culturing human pluripotent and adult stem cells.

Nat Commun 2020 05 19;11(1):2493. Epub 2020 May 19.

Center for Molecular Medicine and Oncode Institute, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG, Utrecht, The Netherlands.

Genetic changes acquired during in vitro culture pose a risk for the successful application of stem cells in regenerative medicine. To assess the genetic risks induced by culturing, we determined all mutations in individual human stem cells by whole genome sequencing. Individual pluripotent, intestinal, and liver stem cells accumulate 3.5 ± 0.5, 7.2 ± 1.1 and 8.3 ± 3.6 base substitutions per population doubling, respectively. The annual in vitro mutation accumulation rate of adult stem cells is nearly 40-fold higher than the in vivo mutation accumulation rate. Mutational signature analysis reveals that in vitro induced mutations are caused by oxidative stress. Reducing oxygen tension in culture lowers the mutational load. We use the mutation rates, spectra, and genomic distribution to model the accumulation of oncogenic mutations during typical in vitro expansion, manipulation or screening experiments using human stem cells. Our study provides empirically defined parameters to assess the mutational risk of stem cell based therapies.
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http://dx.doi.org/10.1038/s41467-020-16323-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7237696PMC
May 2020

Challenges in Establishing Pure Lung Cancer Organoids Limit Their Utility for Personalized Medicine.

Cell Rep 2020 05;31(5):107588

Department of Molecular Oncology and Immunology, the Netherlands Cancer Institute - Antoni van Leeuwenhoek Hospital, 1066 CX Amsterdam, the Netherlands; Oncode Institute, Utrecht, the Netherlands. Electronic address:

Clinical implementation of tumor organoids for personalized medicine requires that pure tumor organoids can be reliably established. Here, we present our experience with organoid cultures from >70 non-small cell lung cancer (NSCLC) samples. We systematically evaluate several methods to identify tumor purity of organoids established from intrapulmonary tumors. Eighty percent of organoids from intrapulmonary lesions have a normal copy number profile, suggesting overgrowth by normal airway organoids (AOs). This is further supported by the failure to detect mutations found in the original tumor in organoids. Histomorphology alone is insufficient to determine tumor purity, but when combined with p63 immunostaining, tumor and normal AOs can be distinguished. Taking into account overgrowth by normal AOs, the establishment rate of pure NSCLC organoids is 17%. Therefore, current methods are insufficient to establish pure NSCLC organoids from intrapulmonary lesions. We discourage their use unless steps are taken to prevent overgrowth by normal AOs.
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http://dx.doi.org/10.1016/j.celrep.2020.107588DOI Listing
May 2020

SARS-CoV-2 productively infects human gut enterocytes.

Science 2020 07 1;369(6499):50-54. Epub 2020 May 1.

Oncode Institute, Hubrecht Institute, Royal Netherlands Academy of Arts and Sciences and University Medical Center, Utrecht, Netherlands.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can cause coronavirus disease 2019 (COVID-19), an influenza-like disease that is primarily thought to infect the lungs with transmission through the respiratory route. However, clinical evidence suggests that the intestine may present another viral target organ. Indeed, the SARS-CoV-2 receptor angiotensin-converting enzyme 2 (ACE2) is highly expressed on differentiated enterocytes. In human small intestinal organoids (hSIOs), enterocytes were readily infected by SARS-CoV and SARS-CoV-2, as demonstrated by confocal and electron microscopy. Enterocytes produced infectious viral particles, whereas messenger RNA expression analysis of hSIOs revealed induction of a generic viral response program. Therefore, the intestinal epithelium supports SARS-CoV-2 replication, and hSIOs serve as an experimental model for coronavirus infection and biology.
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http://dx.doi.org/10.1126/science.abc1669DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7199907PMC
July 2020

Mutational signature in colorectal cancer caused by genotoxic pks E. coli.

Nature 2020 04 27;580(7802):269-273. Epub 2020 Feb 27.

Hubrecht Institute, Royal Netherlands Academy of Arts and Sciences (KNAW) and UMC Utrecht, Utrecht, The Netherlands.

Various species of the intestinal microbiota have been associated with the development of colorectal cancer, but it has not been demonstrated that bacteria have a direct role in the occurrence of oncogenic mutations. Escherichia coli can carry the pathogenicity island pks, which encodes a set of enzymes that synthesize colibactin. This compound is believed to alkylate DNA on adenine residues and induces double-strand breaks in cultured cells. Here we expose human intestinal organoids to genotoxic pks E. coli by repeated luminal injection over five months. Whole-genome sequencing of clonal organoids before and after this exposure revealed a distinct mutational signature that was absent from organoids injected with isogenic pks-mutant bacteria. The same mutational signature was detected in a subset of 5,876 human cancer genomes from two independent cohorts, predominantly in colorectal cancer. Our study describes a distinct mutational signature in colorectal cancer and implies that the underlying mutational process results directly from past exposure to bacteria carrying the colibactin-producing pks pathogenicity island.
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http://dx.doi.org/10.1038/s41586-020-2080-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8142898PMC
April 2020

Impact of panel design and cut-off on tumour mutational burden assessment in metastatic solid tumour samples.

Br J Cancer 2020 03 25;122(7):953-956. Epub 2020 Feb 25.

Department of Medical Oncology, Erasmus MC Cancer Institute, Erasmus MC, Rotterdam, The Netherlands.

Tumour mutational burden (TMB) has emerged as a promising biomarker to predict immune checkpoint inhibitors (ICIs) response in advanced solid cancers. However, harmonisation of TMB reporting by targeted gene panels is lacking, especially in metastatic tumour samples. To address this issue, we used data of 2841 whole-genome sequenced metastatic cancer biopsies to perform an in silico analysis of TMB determined by seven gene panels (FD1CDx, MSK-IMPACT™, Caris Molecular Intelligence, Tempus xT, Oncomine Tumour Mutation Load, NeoTYPE Discovery Profile and CANCERPLEX) compared to exome-based TMB as a golden standard. Misclassification rates declined from up to 30% to <1% when the cut-point for high TMB was increased. Receiver operating characteristic analysis demonstrated that, for correct classification, the cut-point for each gene panel may vary more than 20%. In conclusion, we here demonstrate that a major limitation for the use of gene panels is inter-assay variation and the need for dynamic thresholds to compare TMB outcomes.
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http://dx.doi.org/10.1038/s41416-020-0762-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7109082PMC
March 2020

A deep learning system accurately classifies primary and metastatic cancers using passenger mutation patterns.

Nat Commun 2020 02 5;11(1):728. Epub 2020 Feb 5.

Ontario Institute for Cancer Research, Toronto, ON, M5G0A3, Canada.

In cancer, the primary tumour's organ of origin and histopathology are the strongest determinants of its clinical behaviour, but in 3% of cases a patient presents with a metastatic tumour and no obvious primary. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, we train a deep learning classifier to predict cancer type based on patterns of somatic passenger mutations detected in whole genome sequencing (WGS) of 2606 tumours representing 24 common cancer types produced by the PCAWG Consortium. Our classifier achieves an accuracy of 91% on held-out tumor samples and 88% and 83% respectively on independent primary and metastatic samples, roughly double the accuracy of trained pathologists when presented with a metastatic tumour without knowledge of the primary. Surprisingly, adding information on driver mutations reduced accuracy. Our results have clinical applicability, underscore how patterns of somatic passenger mutations encode the state of the cell of origin, and can inform future strategies to detect the source of circulating tumour DNA.
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http://dx.doi.org/10.1038/s41467-019-13825-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7002586PMC
February 2020

Pancreatic cancer organoids recapitulate disease and allow personalized drug screening.

Proc Natl Acad Sci U S A 2019 Dec 9. Epub 2019 Dec 9.

Oncode Institute, University Medical Center Utrecht, 3584 CX Utrecht, The Netherlands;

We report the derivation of 30 patient-derived organoid lines (PDOs) from tumors arising in the pancreas and distal bile duct. PDOs recapitulate tumor histology and contain genetic alterations typical of pancreatic cancer. In vitro testing of a panel of 76 therapeutic agents revealed sensitivities currently not exploited in the clinic, and underscores the importance of personalized approaches for effective cancer treatment. The PRMT5 inhibitor EZP015556, shown to target (a gene commonly lost in pancreatic cancer)-negative tumors, was validated as such, but also appeared to constitute an effective therapy for a subset of MTAP-positive tumors. Taken together, the work presented here provides a platform to identify novel therapeutics to target pancreatic tumor cells using PDOs.
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http://dx.doi.org/10.1073/pnas.1911273116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6936689PMC
December 2019

Prioritization of genes driving congenital phenotypes of patients with de novo genomic structural variants.

Genome Med 2019 12 4;11(1):79. Epub 2019 Dec 4.

Center for Molecular Medicine and Oncode Institute, University Medical Center Utrecht, 3584 CX, Utrecht, the Netherlands.

Background: Genomic structural variants (SVs) can affect many genes and regulatory elements. Therefore, the molecular mechanisms driving the phenotypes of patients carrying de novo SVs are frequently unknown.

Methods: We applied a combination of systematic experimental and bioinformatic methods to improve the molecular diagnosis of 39 patients with multiple congenital abnormalities and/or intellectual disability harboring apparent de novo SVs, most with an inconclusive diagnosis after regular genetic testing.

Results: In 7 of these cases (18%), whole-genome sequencing analysis revealed disease-relevant complexities of the SVs missed in routine microarray-based analyses. We developed a computational tool to predict the effects on genes directly affected by SVs and on genes indirectly affected likely due to the changes in chromatin organization and impact on regulatory mechanisms. By combining these functional predictions with extensive phenotype information, candidate driver genes were identified in 16/39 (41%) patients. In 8 cases, evidence was found for the involvement of multiple candidate drivers contributing to different parts of the phenotypes. Subsequently, we applied this computational method to two cohorts containing a total of 379 patients with previously detected and classified de novo SVs and identified candidate driver genes in 189 cases (50%), including 40 cases whose SVs were previously not classified as pathogenic. Pathogenic position effects were predicted in 28% of all studied cases with balanced SVs and in 11% of the cases with copy number variants.

Conclusions: These results demonstrate an integrated computational and experimental approach to predict driver genes based on analyses of WGS data with phenotype association and chromatin organization datasets. These analyses nominate new pathogenic loci and have strong potential to improve the molecular diagnosis of patients with de novo SVs.
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http://dx.doi.org/10.1186/s13073-019-0692-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6894143PMC
December 2019

The genomic landscape of metastatic castration-resistant prostate cancers reveals multiple distinct genotypes with potential clinical impact.

Nat Commun 2019 11 20;10(1):5251. Epub 2019 Nov 20.

Department of Medical Oncology, Erasmus MC Cancer Institute, Erasmus University Medical Center Rotterdam, Rotterdam, The Netherlands.

Metastatic castration-resistant prostate cancer (mCRPC) has a highly complex genomic landscape. With the recent development of novel treatments, accurate stratification strategies are needed. Here we present the whole-genome sequencing (WGS) analysis of fresh-frozen metastatic biopsies from 197 mCRPC patients. Using unsupervised clustering based on genomic features, we define eight distinct genomic clusters. We observe potentially clinically relevant genotypes, including microsatellite instability (MSI), homologous recombination deficiency (HRD) enriched with genomic deletions and BRCA2 aberrations, a tandem duplication genotype associated with CDK12 and a chromothripsis-enriched subgroup. Our data suggests that stratification on WGS characteristics may improve identification of MSI, CDK12 and HRD patients. From WGS and ChIP-seq data, we show the potential relevance of recurrent alterations in non-coding regions identified with WGS and highlight the central role of AR signaling in tumor progression. These data underline the potential value of using WGS to accurately stratify mCRPC patients into clinically actionable subgroups.
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http://dx.doi.org/10.1038/s41467-019-13084-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6868175PMC
November 2019
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