bims-fragic 2025-03-16 papers

Issue of 2025–03–16
five papers selected by
Laura Mannarino, Humanitas Research

Front Genet. 2025 ;16 1527884

Exploration of cfDNA landscape in NIPT and clinical utilities of cfDNA based gene expression inference in prenatal diagnostics.

Ruo Jia, Jianjiang Zhu, Feng Zhang, Yangbo Sun, Bin Zhang, Yang Du, Hong Qi.

Cell-free DNA (cfDNA) is a dynamic biomarker reflecting the physiological state of the body. Its unique physical and biochemical properties, inherited from the tissue of origin, enable a wide range of clinical applications. From methylation patterns and fragmentation profiles to genetic variants, cfDNA holds immense potential for diagnosing and monitoring various diseases, including cancer. In this study, we leverage a large collection of non-invasive prenatal testing (NIPT) dataset to explore the genomic landscape of fetal cfDNA, aiming to identify novel biomarkers associated with fetal development and maternal-fetal complications. Our study identifies novel fetal-specific genomic regions, further demonstrating the potential of cfDNA as a versatile biomarker. The prediction model achieved a 100% (12 of 12) positive predictive value (PPV) for hypothyroidism. Whereas for preeclampsia the PPV is much lower (25%, 3 of 12). By establishing a foundation for early hypothyroidism prediction and preeclampsia, we contribute to the expanding applications of NIPT. This approach can be adapted to explore other complex phenotypes and inform biomarker discovery, ultimately advancing maternal-fetal medicine.

Keywords: NIPT; WGS; cfDNA; chromatin; fragmentomics; gene expression; hypothyroidism; preeclampsia

DOI: https://doi.org/10.3389/fgene.2025.1527884

J Mol Neurosci. 2025 Mar 13. 75(1): 34

The Potential of cfDNA as Biomarker: Opportunities and Challenges for Neurodegenerative Diseases.

Şeyma Aydın, Selçuk Özdemir, Ahmet Adıgüzel.

Neurodegenerative disorders, including Alzheimer's disease (AD), Parkinson's disease (PD), multiple sclerosis (MS), and amyotrophic lateral sclerosis (ALS), are characterized by the progressive and gradual degeneration of neurons. The prevalence and rates of these disorders rise significantly with age. As life spans continue to increase in many countries, the number of cases is expected to grow in the foreseeable future. Early and precise diagnosis, along with appropriate surveillance, continues to pose a challenge. The high heterogeneity of neurodegenerative diseases calls for more accurate and definitive biomarkers to improve clinical therapy. Cell-free DNA (cfDNA), including fragmented DNA released into bodily fluids via apoptosis, necrosis, or active secretion, has emerged as a promising non-invasive diagnostic tool for various disorders including neurodegenerative diseases. cfDNA can serve as an indicator of ongoing cellular damage and mortality, including neuronal loss, and may provide valuable insights into disease processes, progression, and therapeutic responses. This review will first cover the key aspects of cfDNA and then examine recent advances in its potential use as a biomarker for neurodegenerative disorders.

Keywords: Cell-free DNA; CfDNA; Circulating biomarkers; Liquid biopsy; Neurodegenerative disorders

DOI: https://doi.org/10.1007/s12031-025-02317-8

Clin Chem. 2025 Mar 11. pii: hvaf011. [Epub ahead of print]

Enrichment of Microbial DNA in Plasma to Improve Pathogen Detection in Sepsis: A Pilot Study.

Eddie G Dominguez, Bradon R McDonald, Haikun Zhang, Michelle D Stephens, Elise C Dietmann, Megan Nedden, Nicole Byington, Sydney Thompson, Mary Junak, Caitlin S Pepperell, Mehreen T Kisat.

BACKGROUND: Diagnosis of sepsis and timely identification of pathogens in critically ill patients remains challenging. Plasma metagenomic sequencing to detect microbial cell-free DNA (mDNA) has shown promise, but low abundance of mDNA in plasma limits sensitivity and necessitates high sequencing depth. mDNA is shorter and more fragmented than human cell-free DNA. Here, we evaluated whether combining single-stranded DNA (ssDNA) sequencing library preparation and size selection can enrich mDNA and improve pathogen detection.
METHODS: We prospectively enrolled 48 trauma patients and collected daily blood samples during the first 10 days of intensive care unit (ICU) admission. For patients with culture-proven infections, we extracted plasma DNA, prepared double-stranded DNA (dsDNA) and ssDNA sequencing libraries, and applied size selection to exclude fragments >110 bp. Following sequencing, we performed taxonomic classification, and evaluated differences in mDNA fractions and in sensitivity for pathogen detection (compared to background noise).
RESULTS: We analyzed 46 plasma samples from 5 patients who developed culture-proven infections, including 17 samples coincident with positive microbial cultures. Size-selected ssDNA libraries showed the total mDNA fraction 204-fold higher on average than conventional dsDNA libraries (P < 0.0001). However, for pathogen-specific DNA (at the genus level), the highest sensitivity was observed in size-selected dsDNA (82%), compared to dsDNA (41%), ssDNA (71%), and size-selected ssDNA (35%) library preparations.
CONCLUSIONS: Our results demonstrate that combining ssDNA library preparation together with fragment size selection improves mDNA yield, potentially reducing sequencing requirements. However, at the genus level, this combination also increases background noise, which limits sensitivity for pathogen detection.

DOI: https://doi.org/10.1093/clinchem/hvaf011

Genome Biol. 2025 Mar 07. 26(1): 49

DAGIP: alleviating cell-free DNA sequencing biases with optimal transport.

Antoine Passemiers, Stefania Tuveri, Tatjana Jatsenko, Adriaan Vanderstichele, Pieter Busschaert, An Coosemans, Dirk Timmerman, Sabine Tejpar, Peter Vandenberghe, Diether Lambrechts, Daniele Raimondi, Joris Robert Vermeesch, Yves Moreau.

Cell-free DNA (cfDNA) is a rich source of biomarkers for various pathophysiological conditions. Preanalytical variables, such as the library preparation protocol or sequencing platform, are major confounders of cfDNA analysis. We present DAGIP, a novel data correction method that builds on optimal transport theory and deep learning, which explicitly corrects for the effect of such preanalytical variables and can infer technical biases. Our method improves cancer detection and copy number alteration analysis by alleviating the sources of variation that are not of biological origin. It also enhances fragmentomic analysis of cfDNA. DAGIP allows the integration of cohorts from different studies.

DOI: https://doi.org/10.1186/s13059-025-03511-y

J Chromatogr B Analyt Technol Biomed Life Sci. 2025 Feb 27. pii: S1570-0232(25)00091-1. [Epub ahead of print]1256 124539

Optimizing nuclease treatment to enhance anion exchange chromatography of HIV-derived virus-like particles.

M S von Elling-Tammen, F Taft, V Thom, J Stitz, S Barbe, A Krause.

Residual host cell chromatin imposes numerous challenges on purifying HIV-derived enveloped virus-like particles (VLPs) using anion-exchange chromatography (AEX). According to FDA guidelines, DNA must be reduced to less than 10 ng per dose at a fragment size of less than 200 bp. To prove the fulfillment of these quality criteria, methods for the qualitative and quantitative analysis of DNA fragments must be applied and adapted to chromatin. DNA and chromatin impede the purification of HIV VLPs with AEX, co-eluting in the same fractions as the VLPs. Although nuclease treatments can be employed, the chromatin structure can shield DNA from nuclease activity. To address these challenges, we adjusted our analytical focus on characterizing the chromatin in our clarified HIV VLP supernatant. We identified two DNA subpopulations: a main large fragment population and a minor population consisting of short fragments below 200 bp. Our findings demonstrated that the larger DNA fragments are the primary issue in our process, as they co-elute with the desired VLPs. To remove the long DNA fragment population, we optimized the nuclease treatment using a Design of Experiment approach to digest the DNA despite the tight chromatin structure. The nucleases Benzonase, Denarase, and M-SAN efficiently digested the DNA removing over 90 % of the DNA. By shredding the long DNA fragments before the AEX step, we successfully separated the HIV VLPs from the remaining short DNA fragments. Combined with nuclease treatment, AEX membrane chromatography offers an efficient single-step purification platform for HIV VLP-based vaccines and other therapeutics.

Keywords: Anion-exchange chromatography; Design of Experiment; Enveloped virus-like particles; Nuclease treatment; Residual chromatin

DOI: https://doi.org/10.1016/j.jchromb.2025.124539