Biology subjects

Jager, M.

Publications and source records attributed to Jager, M..

11 recordsLinked to original sources

bioRxiv2024

Bacterial cell-free DNA profiling reveals co-elevation of multiple bacteria in newborn foals with suspected sepsis

BackgroundSepsis is the leading cause of death in newborn foals. This study investigates whether cell-free DNA (cfDNA) sequencing can enhance bacterial pathogen detection in foals with suspected sepsis and addresses existing knowledge gaps and diagnostic challenges. MethodsWe developed a foal cfDNA sequencing for bacteria identification (cfFBI) workflow, integrating wetlab and computational protocols to detect increased bacterial cfDNA abundance in blood. Specifically, cfFBI focusses on enriching bacterial cfDNA molecules and preventing false positive bacterial identifications. cfFBI was applied to blood samples of 25 hospitalized foals categorized according to the neonatal Systemic Inflammatory Response Syndrome (nSIRS) criteria and 7 healthy foals. ResultscfDNA levels of potential sepsis-causing bacterial genera were elevated in all 11 nSIRS-positive foals compared to healthy foals (n=7), and in 8/11 (72.7%) when compared to both nSIRS-negative (n=4; nSIRS=0) and healthy foals, with multiple genera elevated in 5/11 (45.5%). The total cfDNA concentration, bacterial cfDNA fraction and bacterial diversity were not different between the foal groups. However, nSIRS-positive foals showed significantly different end-motifs in host chromosomal cfDNA, and a decrease in host mitochondrial cfDNA fraction. ConclusionsThis study is the first to demonstrate that cfDNA sequencing in blood samples from newborn foals enables detection of pathogenic bacteria and can help identify novel host-related sepsis biomarkers. The elevated presence of multiple sepsis-causing genera in nSIRS-positive foals and the difference in end-motif, suggests that multibacterial elevation may be more common than previously thought. These findings indicate that cfDNA sequencing holds promise as a future diagnostic tool for identifying sepsis in newborn foals.

pathology↗

bioRxiv2024

Nanopore-based consensus sequencing enables accurate multimodal tumor cell-free DNA profiling

Shallow genome-wide cell-free DNA (cfDNA) sequencing holds great promise for non-invasive cancer monitoring by providing reliable copy number alteration (CNA) and fragmentomic profiles. Single nucleotide variations (SNVs) are, however, much harder to identify with low sequencing depth due to sequencing errors. Here we present Nanopore Rolling Circle Amplification (RCA)-enhanced Consensus Sequencing (NanoRCS), which leverages RCA and consensus calling based on genome-wide long-read nanopore sequencing to enable simultaneous multimodal tumor fraction estimation through SNVs, CNAs, and fragmentomics. Efficacy of NanoRCS is tested on 18 cancer patient samples and 3 healthy controls, demonstrating its ability to reliably detect tumor fractions as low as 0.24%. In vitro experiments confirm that SNV measurements are essential for detecting tumor fractions below 2%. NanoRCS provides the opportunity for cost-effective and rapid processing, which aligns well with clinical needs, particularly in settings where quick and accurate cancer monitoring is essential for personalized treatment strategies.

genomics↗

bioRxiv2021

A novel highly potent inhibitor of TMPRSS2-like proteases blocks SARS-CoV-2 variants of concern and is broadly protective against infection and mortality in mice

The COVID-19 pandemic caused by the SARS-CoV-2 virus remains a global public health crisis. Although widespread vaccination campaigns are underway, their efficacy is reduced against emerging variants of concern (VOCs) 1,2. Development of host-directed therapeutics and prophylactics could limit such resistance and offer urgently needed protection against VOCs 3,4. Attractive pharmacological targets to impede viral entry include type-II transmembrane serine proteases (TTSPs), such as TMPRSS2, whose essential role in the virus lifecycle is responsible for the cleavage and priming of the viral spike protein 5-7. Here, we identify and characterize a small-molecule compound, N-0385, as the most potent inhibitor of TMPRSS2 reported to date. N-0385 exhibited low nanomolar potency and a selectivity index of >106 at inhibiting SARS-CoV-2 infection in human lung cells and in donor-derived colonoids 8. Importantly, N-0385 acted as a broad-spectrum coronavirus inhibitor of two SARS-CoV-2 VOCs, B.1.1.7 and B.1.351. Strikingly, single daily intranasal administration of N-0385 early in infection significantly improved weight loss and clinical outcomes, and yielded 100% survival in the severe K18-human ACE2 transgenic mouse model of SARS-CoV-2 disease. This demonstrates that TTSP-mediated proteolytic maturation of spike is critical for SARS-CoV-2 infection in vivo and suggests that N-0385 provides a novel effective early treatment option against COVID-19 and emerging SARS-CoV-2 VOCs.

microbiology↗

bioRxiv2021

Cancer type classification in liquid biopsies based on sparse mutational profiles enabled through data augmentation and integration

Identifying the cell of origin of cancer is important to guide treatment decisions. However, in patients with cancer of unknown primary (CUP), standard diagnostic tools often fail to identify the primary tumor. As an alternative, machine learning approaches have been proposed to classify the cell of origin based on somatic mutation profiles in the genome of solid tissue biopsies. However, solid biopsies can cause complications and certain tumors are not accessible. A promising alternative would be liquid biopsies, which contain ctDNA originating from the tumor. Problematically, somatic mutation profiles of tumors obtained from liquid biopsies are inherently extremely sparse and current machine learning models fail to perform in this setting. Here we propose an improved machine learning method to deal with the sparse nature of liquid biopsy data. Firstly, we downsample the SNVs in the samples in order to mimic sparse data conditions. Then extensive data augmentation is performed to artificially increase the number of training samples in order to enhance model robustness under sparse data conditions. Finally, we employ data integration to merge information from i) somatic single nucleotide variant (SNV) density across the genome, ii) somatic SNVs in driver genes and iii) trinucleotide motifs. Our adapted method achieves an average accuracy of 0.88 on the data where only 70% of SNVs are retained, which is comparable to an average accuracy of 0.87 with the original model on the full SNV data. Even when only 2% of the data is retained, the average accuracy is 0.65 compared to 0.41 with the original model. The method and results presented here open the way for application of machine learning in the detection of the cell of origin of cancer from sparse liquid biopsy data. Author SummaryThe identification of the cell of origin of cancer is an important step towards more personalized cancer care, but this remains a challenge for patients with cancer of unknown primary (CUP) where the source of the malignancy cannot be identified even after extensive clinical assessment with standard diagnostic methods. Somatic mutation profile-based cell of origin classification has emerged in recent years as a promising alternative diagnostic tool that could circumvent the issues of standard CUP diagnostic. In this approach the somatic mutations are obtained from whole genome sequencing (WGS) of solid tissue biopsies from the tumor. However, needle biopsies from tumor tissue can be challenging, as accessibility to the tumor can be limited and taking a biopsy can cause further complications. For these reasons, liquid biopsies have been proposed as a safer alternative to solid tissue biopsies. Problematically, the circulating tumor DNA fragments available in e.g. blood typically represent a much scarcer tumor source than conventional solid tissue biopsies and therefore liquid biopsies give rise to sparse somatic mutation profiles. Therefore it is crucial to investigate the applicability of sparse somatic mutation profiles in the identification of cell of origin and explore potential improvements of the data analysis and prediction models to overcome sparsity.

bioinformatics↗

bioRxiv2020

Accurate detection of circulating tumor DNA using nanopore consensus sequencing

Levels of circulating tumor DNA (ctDNA) in liquid biopsies may serve as a sensitive biomarker for real-time, minimally-invasive tumor diagnostics and monitoring. However, detecting ctDNA is challenging, as much fewer than 5% of the cell-free DNA in the blood typically originates from the tumor. To detect lowly abundant ctDNA molecules based on somatic variants, extremely sensitive sequencing methods are required. Here, we describe a new technique, CyclomicsSeq, which is based on Oxford Nanopore sequencing of concatenated copies of a single DNA molecule. Consensus calling of the DNA copies increased the base-calling accuracy [~]60x, enabling accurate detection of TP53 mutations at frequencies down to 0.02%. We demonstrate that a TP53-specific CyclomicsSeq assay can be successfully used to monitor tumor burden during treatment for head-and-neck cancer patients. CyclomicsSeq can be applied to any genomic locus and offers an accurate diagnostic liquid biopsy approach that can be implemented in point-of-care clinical workflows.

cancer biology↗

bioRxiv2019

Mutational impact of chronic alcohol use on stem cells in cirrhotic liver

Excessive alcohol consumption increases the risk of developing liver cancer, but the mechanism through which alcohol drives carcinogenesis is as yet unknown. Here, we determined the mutational consequences of chronic alcohol use on the genome of human liver stem cells prior to cancer development. No change in base substitution rate or spectrum could be detected. Analysis of the trunk mutations in an alcohol-related liver tumor by multi-site whole-genome sequencing confirms the absence of specific alcohol-induced mutational signatures driving the development of liver cancer. However, we did identify an enrichment of nonsynonymous base substitutions in cancer genes in stem cells of the cirrhotic livers, such as recurrent nonsense mutations in PTPRK that disturb Epidermal Growth Factor (EGF)-signaling. Our results thus suggest that chronic alcohol use does not contribute to carcinogenesis through altered mutagenicity, but instead induces microenvironment changes which provide a fertile ground for selection of cells with oncogenic mutations.

cancer biology↗

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

The genome of the jellyfish Clytia hemisphaerica and the evolution of the cnidarian life-cycle

Jellyfish (medusae) are a distinctive life-cycle stage of medusozoan cnidarians. They are major marine predators, with integrated neurosensory, muscular and organ systems. The genetic foundations of this complex form are largely unknown. We report the draft genome of the hydrozoan jellyfish Clytia hemisphaerica and use multiple transcriptomes to determine gene use across life-cycle stages. Medusa, planula larva and polyp are each characterised by distinct transcriptome signatures reflecting abrupt life cycle transitions, and all deploy a mixture of phylogenetically old and new genes. Medusa specific transcription factors, including many with bilaterian orthologs, associate with diverse neurosensory structures. Compared to Clytia, the polyp-only hydrozoan Hydra has lost many of the medusa-expressed transcription factors, despite similar overall rates of gene content and sequence evolution. Absence of expression and gene loss among Clytia orthologs of genes patterning the anthozoan aboral pole, secondary axis and endomesoderm support simplification of planulae and polyps in Hydrozoa, including loss of bilateral symmetry. Consequently, although the polyp and planula are generally considered the ancestral cnidarian forms, in Clytia the medusa maximally deploys ancestral cnidarian-bilaterian transcription factor gene complexity.

evolutionary 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↗