scholarly journals Sub-nucleosomal organization in urine cell-free DNA

2019 ◽  
Author(s):  
Havell Markus ◽  
Jun Zhao ◽  
Tania Contente-Cuomo ◽  
Elizabeth Raupach ◽  
Ahuva Odenheimer-Bergman ◽  
...  
Keyword(s):  
Nucleosome Occupancy ◽  
Area Under The Curve ◽  
Wide Distribution ◽  
Cancer Diagnostics ◽  
Cell Of Origin ◽  
Cell Free Dna ◽  
Noninvasive Diagnostics ◽  
Free Dna ◽  
Genome Wide ◽  
Fragmentation Patterns

AbstractCell-free DNA (cfDNA) in urine is a promising analyte for noninvasive diagnostics. However, urine cfDNA is highly fragmented and whether characteristics of these fragments reflect underlying genomic architecture is unknown. Here, we perform comprehensive characterization of fragmentation patterns in urine cfDNA. We show modal size and genome-wide distribution of urine cfDNA fragments are consistent with transient protection from degradation by stable intermediates of nucleosome disassembly. Genome-wide nucleosome occupancy and fragment sizes in urine cfDNA are informative of cell of origin and renal epithelial cells are amongst the highest contributors in urine. Compared to a nucleosome occupancy map based on control urine samples, we observe a higher fraction of fragments with aberrant ends in cancer patients, distinguishing cancer samples with an area under the curve of 0.89. Our results demonstrate sub-nucleosomal organization in urine cfDNA and are proof-of-principle that genome-wide fragmentation analysis of urine cfDNA can enable cancer diagnostics.

scholarly journals Analysis of recurrently protected genomic regions in cell-free DNA found in urine

2021 ◽  
Vol 13 (581) ◽  
pp. eaaz3088
Author(s):  
Havell Markus ◽  
Jun Zhao ◽  
Tania Contente-Cuomo ◽  
Michelle D. Stephens ◽  
Elizabeth Raupach ◽  
...  
Keyword(s):  
Area Under The Curve ◽  
Enzymatic Digestion ◽  
Chromatin Accessibility ◽  
Base Pairs ◽  
Cell Free Dna ◽  
Noninvasive Diagnostics ◽  
Free Dna ◽  
Partial Overlap ◽  
Fragmentation Patterns ◽  
Genomic Regions

Cell-free DNA (cfDNA) in urine is a promising analyte for noninvasive diagnostics. However, urine cfDNA is highly fragmented. Whether characteristics of these fragments reflect underlying genomic architecture is unknown. Here, we characterized fragmentation patterns in urine cfDNA using whole-genome sequencing. Size distribution of urine cfDNA fragments showed multiple strong peaks between 40 and 120 base pairs (bp) with a modal size of 81- and sharp 10-bp periodicity, suggesting transient protection from complete degradation. These properties were robust to preanalytical perturbations, such as at-home collection and delay in processing. Genome-wide sequencing coverage of urine cfDNA fragments revealed recurrently protected regions (RPRs) conserved across individuals, with partial overlap with nucleosome positioning maps inferred from plasma cfDNA. The ends of cfDNA fragments clustered upstream and downstream of RPRs, and nucleotide frequencies of fragment ends indicated enzymatic digestion of urine cfDNA. Compared to plasma, fragmentation patterns in urine cfDNA showed greater correlation with gene expression and chromatin accessibility in epithelial cells of the urinary tract. We determined that tumor-derived urine cfDNA exhibits a higher frequency of aberrant fragments that end within RPRs. By comparing the fraction of aberrant fragments and nucleotide frequencies of fragment ends, we identified urine samples from cancer patients with an area under the curve of 0.89. Our results revealed nonrandom genomic positioning of urine cfDNA fragments and suggested that analysis of fragmentation patterns across recurrently protected genomic loci may serve as a cancer diagnostic.

scholarly journals The effect of pre-analytical and physiological variables on cell-free DNA fragmentation.

2021 ◽  
Author(s):  
Ymke van der Pol ◽  
Norbert Moldovan ◽  
Sandra Verkuijlen ◽  
Jip Ramaker ◽  
Dries Boers ◽  
...  
Keyword(s):  
Biological Properties ◽  
Physiological Factors ◽  
Cell Free Dna ◽  
Physiological Variables ◽  
Processing Times ◽  
Free Dna ◽  
Genome Wide ◽  
Collection Tube ◽  
Fragmentation Patterns ◽  
The Impact

Assays that account for the biological properties and fragmentation of cell-free DNA (cfDNA) can improve the performance of liquid biopsy. However, pre-analytic and physiological differences between individuals on fragmentomic analysis are poorly defined. We analyzed the impact of collection tube, plasma processing time and physiology on the size distribution of cfDNA, their genome-wide representation and sequence diversity at the cfDNA fragment-ends using shallow Whole Genome Sequencing. We observed that using different stabilizing collection tubes, or processing times does not affect the cfDNA fragment sizes, but can impact the genome-wide fragmentation patterns and fragment-end sequences of cfDNA. In addition, beyond differences depending on the gender, the physiological conditions tested between 63 individuals (age, body mass index, use of medication and chronic conditions) minimally influenced the outcome of fragmentomic methods. Our results highlight that fragmentomic approaches have potential for implementation in the clinic, pending clear traceability of analytical and physiological factors.

scholarly journals 5-Hydroxymethylcytosine Signatures in Circulating Cell-Free DNA as Early Warning Biomarkers for COVID-19 Progression and Myocardial Injury

2022 ◽  
Vol 9 ◽  
Author(s):  
Hang-yu Chen ◽  
Xiao-xiao Li ◽  
Chao Li ◽  
Hai-chuan Zhu ◽  
Hong-yan Hou ◽  
...  
Keyword(s):  
Early Warning ◽  
Myocardial Injury ◽  
Area Under The Curve ◽  
Critical Conditions ◽  
Cell Free Dna ◽  
Free Dna ◽  
Genome Wide ◽  
Machine Learning Model ◽  
Warning Model ◽  
Phosphodiesterase 4D

Background: The symptoms of coronavirus disease 2019 (COVID-19) range from moderate to critical conditions, leading to death in some patients, and the early warning indicators of the COVID-19 progression and the occurrence of its serious complications such as myocardial injury are limited.Methods: We carried out a multi-center, prospective cohort study in three hospitals in Wuhan. Genome-wide 5-hydroxymethylcytosine (5hmC) profiles in plasma cell-free DNA (cfDNA) was used to identify risk factors for COVID-19 pneumonia and develop a machine learning model using samples from 53 healthy volunteers, 66 patients with moderate COVID-19, 99 patients with severe COVID-19, and 38 patients with critical COVID-19.Results: Our warning model demonstrated that an area under the curve (AUC) for 5hmC warning moderate patients developed into severe status was 0.81 (95% CI 0.77–0.85) and for severe patients developed into critical status was 0.92 (95% CI 0.89–0.96). We further built a warning model on patients with and without myocardial injury with the AUC of 0.89 (95% CI 0.84–0.95).Conclusion: This is the first study showing the utility of 5hmC as an accurate early warning marker for disease progression and myocardial injury in patients with COVID-19. Our results show that phosphodiesterase 4D and ten-eleven translocation 2 may be important markers in the progression of COVID-19 disease.

scholarly journals FinaleDB: a browser and database of cell-free DNA fragmentation patterns

2020 ◽  
Author(s):  
Haizi Zheng ◽  
Michelle S Zhu ◽  
Yaping Liu
Keyword(s):  
Molecular Mechanisms ◽  
Cell Types ◽  
Cell Free Dna ◽  
Link Type ◽  
Diagnosis And Prognosis ◽  
Dna Database ◽  
Free Dna ◽  
Genome Wide ◽  
Fragmentation Patterns ◽  
Different Cell Types

AbstractSummaryCirculating cell-free DNA (cfDNA) is a promising biomarker for the diagnosis and prognosis of many diseases, including cancer. The genome-wide non-random fragmentation patterns of cfDNA are associated with the nucleosomal protection, epigenetic environment, and gene expression in the cell types that contributed to cfDNA. However, current progress on the development of computational methods and understanding of molecular mechanisms behind cfDNA fragmentation patterns is significantly limited by the controlled-access of cfDNA whole-genome sequencing (WGS) dataset. Here, we present FinaleDB (FragmentatIoN AnaLysis of cEll-free DNA DataBase), a comprehensive database to host thousands of uniformly processed and curated de-identified cfDNA WGS datasets across different pathological conditions. Furthermore, FinaleDB comes with a fragmentation genome browser, from which users can seamlessly integrate thousands of other omics data in different cell types to experience a comprehensive view of both gene-regulatory landscape and cfDNA fragmentation patterns.Availability and implementationFinaleDB service: http://finaledb.research.cchmc.org/ FinaleDB source code: https://github.com/epifluidlab/finaledb_portal and https://github.com/epifluidlab/[email protected]

scholarly journals FinaleDB: a browser and database of cell-free DNA fragmentation patterns

2020 ◽  
Author(s):  
Haizi Zheng ◽  
Michelle S Zhu ◽  
Yaping Liu
Keyword(s):  
Molecular Mechanisms ◽  
Cell Types ◽  
Supplementary Information ◽  
Cell Free Dna ◽  
Diagnosis And Prognosis ◽  
Dna Database ◽  
Free Dna ◽  
Genome Wide ◽  
Fragmentation Patterns ◽  
Different Cell Types

Abstract Summary Circulating cell-free DNA (cfDNA) is a promising biomarker for the diagnosis and prognosis of many diseases, including cancer. The genome-wide non-random fragmentation patterns of cfDNA are associated with the nucleosomal protection, epigenetic environment, and gene expression in the cell types that contributed to cfDNA. However, current progress on the development of computational methods and understanding of molecular mechanisms behind cfDNA fragmentation patterns is significantly limited by the controlled-access of cfDNA whole-genome sequencing (WGS) dataset. Here, we present FinaleDB (FragmentatIoN AnaLysis of cEll-free DNA DataBase), a comprehensive database to host thousands of uniformly processed and curated de-identified cfDNA WGS datasets across different pathological conditions. Furthermore, FinaleDB comes with a fragmentation genome browser, from which users can seamlessly integrate thousands of other omics data in different cell types to experience a comprehensive view of both gene-regulatory landscape and cfDNA fragmentation patterns. Availability and implementation FinaleDB service: http://finaledb.research.cchmc.org/. FinaleDB source code: https://github.com/epifluidlab/finaledb_portal, https://github.com/epifluidlab/finaledb_workflow Supplementary information Supplementary data are available at Bioinformatics online.

Genome-wide cell-free DNA fragmentation profiling for early cancer detection.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. 3018-3018
Author(s):  
Alessandro Leal ◽  
Stephen Cristiano ◽  
Jillian Phallen ◽  
Jacob Fiksel ◽  
Vilmos Adleff ◽  
...  
Keyword(s):  
Dna Fragmentation ◽  
White Blood Cells ◽  
Targeted Sequencing ◽  
Healthy Individuals ◽  
Cell Free Dna ◽  
Patients With Cancer ◽  
Molecular Features ◽  
Free Dna ◽  
Genome Wide ◽  
Fragmentation Patterns

3018 Background: Analyses of cell-free DNA (cfDNA) in the blood provide a noninvasive diagnostic avenue for patients with cancer. However, cfDNA analyses have largely focused on targeted sequencing of specific genes, and the characteristics of the origins and molecular features of cfDNA are poorly understood. We developed an ultrasensitive approach that allows simultaneous examination of a large number of abnormalities in cfDNA through genome-wide analysis of fragmentation patterns. Methods: We used a machine learning model to examined cfDNA fragmentation profiles of 236 patients with largely localized breast, colorectal, lung, ovarian, pancreatic, gastric, or bile duct cancer and 245 healthy individuals. Estimation of performance was determined by ten-fold cross validation repeated ten times. Results: cfDNA profiles of healthy individuals reflected nucleosomal patterns of white blood cells, while patients with cancer had altered fragmentation patterns. The degree of abnormality in fragmentation profiles during therapy closely matched levels of mutant allele fractions in cfDNA as determined using ultra-deep targeted sequencing. The sensitivity of detection ranged from 57% to > 99% among the seven cancer types at 98% specificity, with an overall AUC of 0.94. Fragmentation profiles could be used to identify the tissue of origin of the cancers to a limited number of sites in 75% of cases. Combining our approach with mutation-based cfDNA analyses detected 91% of cancer patients. Conclusions: This effort is the first study to demonstrate genome-wide cell-free DNA fragmentation abnormalities in patients with cancer. Results of these analyses highlight important properties of cfDNA and provide a facile approach for screening, early detection, and monitoring of human cancer.

scholarly journals Detecting cell-of-origin and cancer-specific features of cell-free DNA with Nanopore sequencing

2021 ◽  
Author(s):  
Efrat Katsman ◽  
Shari Orlanski ◽  
Filippo Martignano ◽  
Amir Eden ◽  
Iacopo Petrini ◽  
...  
Keyword(s):  
Circulating Tumor Dna ◽  
Cell Of Origin ◽  
Cell Free Dna ◽  
Bisulfite Treatment ◽  
Free Dna ◽  
Oxford Nanopore ◽  
Genomic Regulation ◽  
Fragmentation Patterns ◽  
Oxford Nanopore Technologies ◽  
Processing Steps

DNA methylation (5mC) is a promising biomarker for detecting circulating tumor DNA (ctDNA), providing information on a cell's genomic regulation, developmental lineage, and molecular age. Sequencing assays for detecting ctDNA methylation involve pre-processing steps such as immunoprecipitation, enzymatic treatment, or the most common method, sodium bisulfite treatment. These steps add complexity and time that pose a challenge for clinical labs, and bisulfite treatment in particular degrades input DNA and can result in loss of informative ctDNA fragmentation patterns. In this feasibility study, we demonstrate that whole genome sequencing of circulating cell-free DNA using conventional Oxford Nanopore Technologies (ONT) sequencing can accurately detect cell-of-origin and cancer-specific 5mC changes while preserving important fragmentomic information. The simplicity of this approach makes it attractive as a liquid biopsy assay for cancer as well as non-cancer applications in emergency medicine.

scholarly journals Aging-associated distinctive DNA methylation changes of LINE-1 retrotransposons in pure cell-free DNA from human blood

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Wardah Mahmood ◽  
Lars Erichsen ◽  
Pauline Ott ◽  
Wolfgang A. Schulz ◽  
Johannes C. Fischer ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Cell Free Dna ◽  
The Past ◽  
Free Dna ◽  
Genome Wide ◽  
Cancerous Cell ◽  
Pure Cell ◽  
Epigenetic Biomarker ◽  
Methylation Patterns ◽  
Cancerous Cells

AbstractLINE-1 hypomethylation of cell-free DNA has been described as an epigenetic biomarker of human aging. However, in the past, insufficient differentiation between cellular and cell-free DNA may have confounded analyses of genome-wide methylation levels in aging cells. Here we present a new methodological strategy to properly and unambiguously extract DNA methylation patterns of repetitive, as well as single genetic loci from pure cell-free DNA from peripheral blood. Since this nucleic acid fraction originates mainly in apoptotic, senescent and cancerous cells, this approach allows efficient analysis of aged and cancerous cell-specific DNA methylation patterns for diagnostic and prognostic purposes. Using this methodology, we observe a significant age-associated erosion of LINE-1 methylation in cfDNA suggesting that the threshold of hypomethylation sufficient for relevant LINE-1 activation and consequential harmful retrotransposition might be reached at higher age. We speculate that this process might contribute to making aging the main risk factor for many cancers.

scholarly journals The use of plasma donor-derived, cell-free DNA to monitor acute rejection after kidney transplantation

2019 ◽  
Vol 35 (4) ◽  
pp. 714-721 ◽  
Author(s):  
Els M Gielis ◽  
Kristien J Ledeganck ◽  
Amélie Dendooven ◽  
Pieter Meysman ◽  
Charlie Beirnaert ◽  
...  
Keyword(s):  
Acute Rejection ◽  
Serum Creatinine ◽  
Characteristic Curve ◽  
Area Under The Curve ◽  
Threshold Value ◽  
Nucleotide Polymorphisms ◽  
Kidney Transplant Recipients ◽  
Cell Free Dna ◽  
Free Dna ◽  
Set Up

Abstract Background After transplantation, cell-free deoxyribonucleic acid (DNA) derived from the donor organ (ddcfDNA) can be detected in the recipient’s circulation. We aimed to investigate the role of plasma ddcfDNA as biomarker for acute kidney rejection. Methods From 107 kidney transplant recipients, plasma samples were collected longitudinally after transplantation (Day 1 to 3 months) within a multicentre set-up. Cell-free DNA from the donor was quantified in plasma as a fraction of the total cell-free DNA by next generation sequencing using a targeted, multiplex polymerase chain reaction-based method for the analysis of single nucleotide polymorphisms. Results Increases of the ddcfDNA% above a threshold value of 0.88% were significantly associated with the occurrence of episodes of acute rejection (P = 0.017), acute tubular necrosis (P = 0.011) and acute pyelonephritis (P = 0.032). A receiver operating characteristic curve analysis revealed an equal area under the curve of the ddcfDNA% and serum creatinine of 0.64 for the diagnosis of acute rejection. Conclusions Although increases in plasma ddcfDNA% are associated with graft injury, plasma ddcfDNA does not outperform the diagnostic capacity of the serum creatinine in the diagnosis of acute rejection.

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