Biology subjects

Cuppen, E.

Publications and source records attributed to Cuppen, E..

29 records · Page 2Linked to original sources

bioRxiv2019

5-Fluorouracil treatment induces characteristic T>G mutations in human cancer

5-Fluorouracil (5-FU) is a chemotherapeutic drug component that is commonly used for the treatment of solid cancers. It is proposed that 5-FU possesses anticancer properties via the interference with nucleotide synthesis and incorporation into DNA. As both mechanisms may have a mutational impact on both surviving tumor and healthy cells, we treated intestinal organoids with 5-FU followed by whole genome sequencing analysis and uncovered a highly characteristic mutational pattern that is dominated by T>G substitutions in a CTT context. Analysis of tumor whole genome sequencing data confirmed that this signature can also be identified in vivo in colorectal and breast cancer patients that have undergone treatment with 5-FU. We also found that more 5-FU mutations are induced in TP53 null backgrounds which may be of clinical relevance. Taken together, our results demonstrate that 5-FU is mutagenic and may drive tumor evolution and increase the risk of secondary malignancies. Furthermore, the identified signature shows a strong resemblance to COSMIC signature 17, the hallmark signature of treatment-naive esophageal and gastric tumors, which indicates that distinct endogenous and exogenous triggers can converge onto highly similar mutational signatures.

cancer biology↗

bioRxiv2019

Sperm DNA damage causes genomic instability in early embryonic development

Genomic instability is common in early embryo development, but the underlying causes are largely unknown. Here we examined the consequences of sperm DNA damage on the embryonic genome by single-cell genome sequencing of individual blastomeres from bovine embryos produced with sperm damaged by radiation. Sperm DNA damage caused fragmentation of chromosomes and segregation errors such as heterogoneic cell divisions yielding a broad spectrum of genomic aberrations that are similar to those frequently found in human embryos. The mosaic aneuploidies, mixoploidy, uniparental disomies and de novo structural variation induced upon sperm DNA damage may compromise health and lead to rare genomic disorders when embryos escape developmental arrest.\n\nOne Sentence SummaryDNA damage in sperm cells leads to genomic defects in embryos

developmental biology↗

bioRxiv2019

The genomic landscape of metastatic castration-resistant prostate cancers using whole genome sequencing reveals multiple distinct genotypes with potential clinical impact

Here we present whole-genome sequencing (WGS) analysis of fresh-frozen metastatic biopsies from 197 castration-resistant prostate cancer patients. Using hierarchical unsupervised clustering based on genomic aberrations only, we defined eight different clusters. We detected four distinct and potentially clinically relevant genotypes harboring unique genomic features, including: 1) Microsatellite Instability; 2) Homologous Recombination Deficiency (HRD) with enriched genomic deletions and BRCA2 aberrations; 3) tandem duplication phenotype associated with biallelic CDK12 mutations; and 4) a subgroup enriched for chromothripsis events. Our data suggest that classifying patients using WGS characteristics may improve classification of HRD patients. Moreover, we confirmed that important regulators of AR-mediated signaling are located in non-coding regions. Using ChIP sequencing data, we showed that the amplified AR and MYC promoter regions contain open chromatin and bind AR, suggesting a role in AR mediated biology. Thus, high-resolution WGS may be used to improve patient stratification.

cancer biology↗

bioRxiv2019

Tumor mutational landscape is a record of the pre-malignant state

Chromatin structure has a major influence on the cell-specific density of somatic mutations along the cancer genome. Here, we present a pan-cancer study in which we searched for the putative cancer cell-of-origin of 2,550 whole genomes, representing 32 cancer types by matching their mutational landscape to the regional patterns of chromatin modifications ascertained in 104 normal tissue types. We found that, in almost all cancer types, the cell-of-origin can be predicted solely from their DNA sequences. Our analysis validated the hypothesis that high-grade serous ovarian cancer originates in the fallopian tube and identified distinct origins of breast cancer subtypes. We also demonstrated that the technique is equally capable of identifying the cell-of-origin for a series of 2,044 metastatic samples from 22 of the tumor types available as primaries. Moreover, cancer drivers, whether inherited or acquired, reside in active chromatin regions in the respective cell-of-origin. Taken together, our findings highlight that many somatic mutations accumulate while the chromatin structure of the cell-of-origin is maintained and that this historical record, captured in the DNA, can be used to identify the often elusive cancer cell-of-origin.

genomics↗

bioRxiv2018

Early divergence of mutational mechanisms drives genetic heterogeneity of fetal tissues

A developing human fetus needs to balance rapid cellular expansion with maintaining genomic stability. Here, we accurately quantified and characterized somatic mutation accumulation in fetal tissues by analyzing individual stem cells from human fetal liver and intestine. Fetal mutation rates were ~5-fold higher than in tissue-matched adult stem cells. The mutational landscape of fetal intestinal stem cells resembled that of adult intestinal stem cells, while the mutation spectrum of fetal liver stem cells is distinct from stem cells of the fetal intestine and the adult liver. Our analyses indicate that variation in mutational mechanisms, including oxidative stress and spontaneous deamination of methylated cytosines, contribute to the observed divergence in mutation accumulation patterns and drive genetic mosaicism in humans.\n\nOne Sentence SummaryLiver and intestinal cells accumulate elevated amounts and diverged types of somatic DNA mutations during early human fetal development

genomics↗

bioRxiv2018

Mutational impact of culturing human pluripotent and adult stem cells

Genetic changes acquired during in vitro culture pose a potential risk for the successful application of stem cells in regenerative medicine. To assess mutation accumulation risks induced by culturing, we determined genetic aberrations in individual human induced pluripotent stem cells (iPS cells) and adult stem cells (ASCs) by whole genome sequencing analyses. Individual iPS cells, intestinal ASCs and liver ASCs accumulated 3.5{+/-}0.5, 7.2{+/-}1.0 and 8.4{+/-}3.6 base substitutions per population doubling, respectively. The annual in vitro mutation accumulation rate of ASCs adds up to [~]1600 base pair substitutions, which is [~]40-fold higher than the in vivo rate of [~]40 base pair substitutions per year. Mutational analysis revealed a distinct in vitro induced mutational signature that is irrespective of stem cell type and distinct from the in vivo mutational signature. This in vitro signature is characterized by C to A changes that have previously been linked to oxidative stress conditions. Additionally, we observed stem cell-specific mutational signatures and differences in transcriptional strand bias, indicating differential activity of DNA repair mechanisms between stem cell types in culture. We demonstrate that the empirically defined mutation rates, spectra, and genomic distribution enable risk assessment by modelling the accumulation of specific oncogenic mutations during typical in vitro expansion, manipulation or screening experiments using human stem cells. Taken together, we have here for the first time accurately quantified and characterized in vitro mutation accumulation in human iPS cells and ASCs in a direct comparison. These results provide insights for further optimization of culture conditions for safe in vivo utilization of these cell types for regenerative purposes.

genomics↗

bioRxiv2018

Pan-cancer whole genome analyses of metastatic solid tumors

Metastatic cancer is one of the major causes of death and is associated with poor treatment efficiency. A better understanding of the characteristics of late stage cancer is required to help tailor personalised treatment, reduce overtreatment and improve outcomes. Here we describe the largest pan-cancer study of metastatic solid tumor genomes, including 2,520 whole genome-sequenced tumor-normal pairs, analyzed at a median depth of 106x and 38x respectively, and surveying over 70 million somatic variants. Metastatic lesions were found to be very diverse, with mutation characteristics reflecting those of the primary tumor types, although with high rates of whole genome duplication events (56%). Metastatic lesions are relatively homogeneous with the vast majority (96%) of driver mutations being clonal and up to 80% of tumor suppressor genes bi-allelically inactivated through different mutational mechanisms. For 62% of all patients, genetic variants that may be associated with outcome of approved or experimental therapies were detected. These actionable events were distributed across various mutation types underlining the importance of comprehensive genomic tumor profiling for cancer precision medicine.

cancer biology↗

bioRxiv2018

Long-term expanding human airway organoids for disease modelling.

Organoids are self-organizing 3D structures grown from stem cells that recapitulate essential aspects of organ structure and function. Here we describe a method to establish long-term-expanding human airway organoids from broncho-alveolar biopsies or lavage material. The pseudostratified airway organoid epithelium consists of basal cells, functional multi-ciliated cells, mucus-producing goblet cells, and CC10-secreting club cells. Airway organoids derived from cystic fibrosis (CF) patients allow assessment of CFTR function in an organoid swelling assay. Organoid culture conditions also allow gene editing as well as the derivation of various types of lung cancer organoids. Respiratory syncytial virus (RSV) infection recapitulated central disease features and dramatically increases organoid cell motility, found to be driven by the non-structural viral NS2 protein. We conclude that human airway organoids represent versatile models for the in vitro study of hereditary, malignant, and infectious pulmonary disease.

cell biology↗

bioRxiv2017

Deficiency of global genome nucleotide excision repair explains 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. Tumors deficient in NER are more sensitive to cisplatin treatment. Characterization of the mutational consequences of NER-deficiency may therefore provide important diagnostic opportunities. Here, we analyzed the somatic mutational profiles of adult stem cells (ASCs) from NER-deficient Ercc1-/{Delta} mice, using whole-genome sequencing analysis of clonally derived organoid cultures. Our results indicate that NER-deficiency increases the base substitution load in liver, but not in small intestinal ASCs, which coincides with a tissue-specific aging-pathology observed in these mice. The mutational landscape changes as a result of NER-deficiency in ASCs of both tissues and shows an increased contribution of Signature 8 mutations, which is a pattern with unknown etiology that is recurrently observed in various cancer types. The scattered genomic distribution of the acquired base substitutions indicates that deficiency of global-genome NER (GG-NER) is responsible for the altered mutational landscape. In line with this, we observed 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 may therefore serve as a biomarker for NER-deficiency and could improve personalized cancer treatment strategies.

molecular biology↗

bioRxiv2017

A system-wide approach to monitor responses to synergistic BRAF and EGFR inhibition in colorectal cancer cells

Intrinsic and/or acquired resistance represents one of the great challenges in targeted cancer therapy. A deeper understanding of the molecular biology of cancer has resulted in more efficient strategies, where one or multiple drugs are adopted in novel therapies to tackle resistance. This beneficial effect of using combination treatments has also been observed in colorectal cancer patients harboring the BRAF(V600E) mutation, whereby dual inhibition of BRAF(V600E) and EGFR increases antitumor activity. Notwithstanding this success, it is not clear whether this combination treatment is the only or most effective treatment to block intrinsic resistance to BRAF inhibitors. Here, we investigate molecular responses upon single and multi-target treatments, over time, using BRAF(V600E) mutant colorectal cancer cells as a model system. Through integration of transcriptomic, proteomic and phosphoproteomics data we obtain a comprehensive overview, revealing both known and novel responses. We primarily observe widespread upregulation of receptors tyrosine kinases and metabolic pathways upon BRAF inhibition. These findings point to mechanisms by which the drug-treated cells switch energy sources and enter a quiescent-like state as a defensive response, while additionally reactivating the MAPK pathway.

cancer biology↗

bioRxiv2017

Mapping And Phasing Of Structural Variation In Patient Genomes Using Nanopore Sequencing

Structural genomic variants form a common type of genetic alteration underlying human genetic disease and phenotypic variation. Despite major improvements in genome sequencing technology and data analysis, the detection of structural variants still poses challenges, particularly when variants are of high complexity. Emerging long-read single-molecule sequencing technologies provide new opportunities for detection of structural variants. Here, we demonstrate sequencing of the genomes of two patients with congenital abnormalities using the ONT MinION at 11x and 16x mean coverage, respectively. We developed a bioinformatic pipeline - NanoSV - to efficiently map genomic structural variants (SVs) from the long-read data. We demonstrate that the nanopore data are superior to corresponding short-read data with regard to detection of de novo rearrangements originating from complex chromothripsis events in the patients. Additionally, genome-wide surveillance of SVs, revealed 3,253 (33%) novel variants that were missed in short-read data of the same sample, the majority of which are duplications < 200bp in size. Long sequencing reads enabled efficient phasing of genetic variations, allowing the construction of genome-wide maps of phased SVs and SNVs. We employed read-based phasing to show that all de novo chromothripsis breakpoints occurred on paternal chromosomes and we resolved the long-range structure of the chromothripsis. This work demonstrates the value of long-read sequencing for screening whole genomes of patients for complex structural variants.

genomics↗