Journal of Molecular Diagnostics最新文献

{"title":"Validation of a Modular Gene Expression Assay for Risk Stratification and Subtyping Lymphomas.","authors":"Peter J B Sabatini, Josh Bridgers, Shujun Huang, Tong Zhang, Clare Sheen, Tracy Stockley, Robert Kridel, Ian Bosdet, Marco A Marra, Christian Steidl, David W Scott, Aly Karsan","doi":"10.1016/j.jmoldx.2025.09.004","DOIUrl":"https://doi.org/10.1016/j.jmoldx.2025.09.004","url":null,"abstract":"<p><p>Gene expression signatures are important for classifying lymphoid malignancies, although routine diagnostic workflow predominantly use immunohistochemical staining and fluorescence in situ hybridization. These traditional methods are labor-intensive and may misclassify the underlying oncogenic signatures leading to inaccurate prognostication. To address this, an RNA expression panel was developed, the Lymphoma Expression Analysis (LExA120) 120 gene expression panel, using the NanoString platform for rapid, modular analysis of various lymphoma subtypes. The LExA120 panel targets 95 genes and 25 housekeeping genes to evaluate aggressive B-cell lymphomas, including diffuse large B-cell lymphoma cell-of-origin, dark zone, and primary mediastinal large B-cell lymphoma signatures, Epstein-Barr virus (EBV) status and a classical Hodgkin lymphoma post-transplant risk. Fifty-four formalin-fixed paraffin-embedded tissue samples were tested with known diagnoses and 51 samples with known EBV status. The panel demonstrated high concordance with previously validated methods according to Pearson correlation coefficients of the signature scores. The assay also showed high reproducibility in repeated tests and across different clinical laboratories. Our study confirmed the panel's ability to stratify EBV-positive and EBV-negative lymphomas with high diagnostic certainty. Although EBER in situ hybridization confirmation was needed in approximately 12% of cases, synergizing with traditional techniques may facilitate more rapid and cost-effective diagnoses. The LExA120 panel offers a multiplexed approach to lymphoma classification, enhancing the efficiency and accuracy for subtyping lymphomas.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-10-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145281585","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Enhanced detection of splice altering variants in hematologic malignancies using targeted RNA-seq data.","authors":"Muneeza Maqsood, John Toubia, Carol Wadham, Naranie Shanmuganathan, Nur Hezrin Shahrin, Adelina Fernandes, Joe McConnell, Verity Saunders, Dominik Kaczorowski, Rosalie R Kenyon, Ming Lin, Timothy P Hughes, Chung Hoow Kok, Susan Branford","doi":"10.1016/j.jmoldx.2025.09.003","DOIUrl":"https://doi.org/10.1016/j.jmoldx.2025.09.003","url":null,"abstract":"<p><p>RNA-based targeted sequencing aids the detection of several types of variants in hematologic and other malignancies, including splice-altering variants. However, accurately identifying clinically relevant mis-splicing events remains challenging due to the inherent complexity of the human transcriptome and the high prevalence of false-positive splice junctions in deep RNA sequencing data. To address these challenges, SpliceChaser and BreakChaser were developed, which are bioinformatics tools designed to enhance the detection and characterization of relevant splice altering events. SpliceChaser improves the identification of clinically relevant atypical splicing by analyzing read length diversity within flanking sequences of the mapped reads around the splice junctions. BreakChaser processes soft-clipped sequences and alignment anomalies to enhance the detection of targeted deletion breakpoints associated with atypical splice isoforms generated from intrachromosomal gene deletions. These tools were developed and validated using a cohort of over 1,400 RNA-seq samples from patients with chronic myeloid leukemia (CML). Collectively, SpliceChaser and BreakChaser achieved a positive percentage agreement of 98% and a positive predictive value of 91% for the detection of clinically relevant atypical splice altering variants or gene deletions in the targeted regions. By integrating splicing and breakpoint detection with robust filtering strategies, these tools facilitate precise identification of clinically relevant variants, paving the way for improved diagnostics and therapeutic strategies in CML and other malignancies.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-10-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145281565","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Comparative performance of a targeted next-generation sequencing assay against multiplexed digital PCR assays in detecting ERBB2, ESR1, and PIK3CA mutations in plasma circulating cell-free DNA from liquid biopsies.","authors":"Julien Corné, Florence Godey, Angelina Legros, Laurent Castéra, Sophie Krieger, Mathilde Cherel, Agathe Cochet, Fanny Le Du, Héloïse Bourien, Antoine Deleuze, Laurence Crouzet, Christophe Perrin, Claudia Lefeuvre-Plesse, Véronique Diéras, Thibault De la Motte Rouge","doi":"10.1016/j.jmoldx.2025.08.012","DOIUrl":"https://doi.org/10.1016/j.jmoldx.2025.08.012","url":null,"abstract":"<p><p>Analyzing somatic mutations in liquid biopsies poses a real challenge in treating patients with breast cancer. Because of the high sensitivity required to detect circulating tumor DNA, which may be present at very low levels, digital PCR analysis seems highly appropriate. However, new targeted next-generation sequencing solutions are now available, enabling highly sensitive multi-gene analysis that could benefit patients. This study compared the analytical performance of multiplex digital PCR and targeted next-generation sequencing in detecting somatic erb-b2 receptor tyrosine kinase 2 (ERBB2), estrogen receptor 1 (ESR1), and phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) mutations in a series of 32 plasma samples from patients with metastatic breast cancer. Forty-four mutations were detected, with an overall concordance of 95% (90/95) and a high degree of correlation (R² = 0.9786). Interestingly, two ESR1 mutations detected in multiplex drop-off digital PCR were also detected by targeted next-generation sequencing with comparable mutant allele frequencies, enabling the identification of these specific variants (p.D538N and p.536LYD>P). Moreover, an additional PIK3CA mutation (p.P539R) was first detected using targeted next-generation sequencing and later confirmed with a newly designed digital PCR assay. While more expensive than multiplex digital PCR, these new types of small targeted next-generation sequencing gene panels could provide a rapid answer to a specific clinical question with a ready-to-use solution, which could benefit patients.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-10-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145276489","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Clinical Utility of Multitissue Genomic Arrays in Diagnosing Pigmentary Mosaicism Associated with Neurodevelopmental Delay.","authors":"Yanca Gasparini Oliveira, Marilia Moreira Montenegro, Vanessa Tavares Almeida, Eder Alencar Moura, Amom Mendes Nascimento, Gleyson Francisco da Silva Carvalho, Evelin Aline Zanardo, Samar Nasser Chehimi, Beatriz Martins Wolff, Lucas Liro Vieira, Mariana Ribeiro Costa Siemann, Rafaela da Silva Mendes, Lissandro de Sousa Rolim, Karina Marinho Nascimento, Chong Ae Kim, Leslie Domenici Kulikowski","doi":"10.1016/j.jmoldx.2025.09.002","DOIUrl":"10.1016/j.jmoldx.2025.09.002","url":null,"abstract":"<p><p>Genomic mosaicism is underdiagnosed owing to its variable tissue distribution and the limitations of single-tissue testing. Cytogenomic techniques applied across multiple tissues can uncover clinically actionable variants and clarify genotype-phenotype relationships. DNA from 21 patients (14 females and 7 males) with pigmentary mosaicism and global developmental delay was analyzed. Each patient underwent G-band karyotyping and array (Infinium CytoSNP-850K) on peripheral blood, skin fibroblasts, and buccal mucosa samples. Pathogenic or likely pathogenic variants were found in 13 of 21 patients (62%). Of these 13 patients, 10 (77%) exhibited mosaicism, with variant allele fractions as low as 15%. On the basis of tissue distribution, 3 cases were classified as germline events and 10 as somatic mosaicism. Patients with positive findings were subdivided into: i) mosaic numerical chromosomal alterations (n = 6), ii) pathogenic copy number variations (n = 5), and iii) structural rearrangements (n = 2). Notably, several mosaic variants-particularly aneuploidies-were detected exclusively in fibroblast DNA, underlining the added diagnostic yield of multitissue sampling. In this cohort, a multitissue cytogenomic approach achieved a 62% overall diagnostic rate and identified mosaicism in 77% of positive cases. These results support the routine incorporation of genomic arrays with multisample analysis into diagnostic workflows for rare developmental disorders, enhancing detection sensitivity and enabling precise genotype-phenotype correlations.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-10-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145240189","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Developing Consensus for a More Provider-Friendly Next-Generation Sequencing Molecular Biomarker Report: A Joint Consensus Recommendation of the Association for Molecular Pathology and College of American Pathologists.","authors":"Jane Gibson, Hanadi El Achi, Dana Altenburger, Noah Brown, Amy S Clark, Joshua Coleman, Rajyasree Emmadi, Amber M Fussell, Meera Hameed, Danielle Jordan, Jennifer Laudadio, Anthony Provenzano, Robyn L Temple-Smolkin, Christopher G Suciu","doi":"10.1016/j.jmoldx.2025.08.011","DOIUrl":"10.1016/j.jmoldx.2025.08.011","url":null,"abstract":"<p><p>Despite the increasing availability of next-generation sequencing (NGS) gene panel analysis for cancers, published reports suggest underutilization of testing, citing the shortage of credentialed professionals available to assist with the interpretation of test results among the key barriers. Obtaining a multidisciplinary consensus regarding a shared best practice NGS molecular biomarker reporting section template may facilitate introduction and/or increased testing for institutions that adopt similar report structures by improving report effectiveness and efficiency for both health care providers and laboratory professionals, leading to improved patient care. To address this challenge, the Association for Molecular Pathology convened a multidisciplinary collaborative expert working group to identify and utilize best practices from current reporting guidelines and approaches to develop an NGS biomarker report template to optimally present complex molecular profiling information for efficient use by oncologists and other health care providers. Seventeen non-small-cell lung cancer NGS biomarker reports from public, private, and academic laboratories were reviewed, and specific components (eg, report length, color use, formatting, presentation order of information, specific information included or omitted, tables, and figures) were assessed for their ability to be considered provider friendly. Based on this review, public and stakeholder input, available literature, and cumulative professional experience of the working group members, a guideline-concordant reporting template was developed based on expert opinion consensus and made freely available online with planned implementation assessment.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-09-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145187375","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Analysis of the Fragmentation and Integrity of Urine Cell-Free DNA as a Diagnostic and Staging Biomarker for Bladder Cancer.","authors":"Raquel Herranz, Julia Oto, Emma Plana, Javier Pérez-Ardavín, Patricia Verger, Manuel Martínez-Sarmiento, César D Vera-Donoso, Pilar Medina","doi":"10.1016/j.jmoldx.2025.08.010","DOIUrl":"10.1016/j.jmoldx.2025.08.010","url":null,"abstract":"<p><p>Bladder cancer (BC) is a lethal urological malignancy, with current diagnostic and follow-up methods being invasive and costly. Cell-free DNA (cfDNA) in liquid biopsies has shown promise in cancer diagnostics, but its fragmentation and integrity in urine remain underexplored in BC, becoming the aim of this study. cfDNA was isolated from the urine of 156 patients with BC of most stages and 79 matched controls without renal or bladder conditions. The amount of a large (>250-bp) and a nested small (<125-bp) fragment of ACTB, AR, MYC, BCAS1, and STOX1 was quantified by quantitative real-time PCR. Fragmentation and integrity (ratio of large/small) were analyzed with ordinal logistic regression. The increase in the ratio of large/small ACTB fragments and the small fragments of AR and MYC may represent a valuable tool to diagnose and stage BC when classified as both non-muscle-invasive and muscle-invasive BC or considering grades and stages separately. The small fragment of MYC, leading the effect observed, displayed a valuable diagnostic capacity (area under the curve = 0.7221; 95% CI, 0.6527-0.7915; P < 0.0001; sensitivity = 50%; specificity = 95%), particularly for muscle-invasive BC (area under the curve = 0.8098; 95% CI, 0.6674-0.9523; P < 0.0001; sensitivity = 70%; specificity = 97%). Herein, the analysis of urine cfDNA fragmentation and integrity of these surrogate markers is proposed as noninvasive biomarkers to diagnose and stage BC. Once validated, the proposed biomarkers could improve patient management by reinforcing or substituting current invasive and expensive techniques.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-09-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145187330","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluation of Long-Read Genome Sequencing for Genomic Profiling of Myeloid Cancers.","authors":"Haley J Abel, Mohamed Mahgoub, Nidhi Davarapalli, Rohan Kodgule, Christopher A Miller, Robert S Fulton, Catrina Fronick, Christopher Markovic, Sharon Heath, Jacqueline E Payton, Meagan A Jacoby, Daniel C Link, Matthew J Walter, Eric J Duncavage, Timothy J Ley, David H Spencer","doi":"10.1016/j.jmoldx.2025.09.001","DOIUrl":"10.1016/j.jmoldx.2025.09.001","url":null,"abstract":"<p><p>Whole-genome sequencing (WGS) is a comprehensive approach for the genomic evaluation of acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS). We recently described a streamlined tumor-only WGS assay (ChromoSeq) that uses Illumina short-read sequencing with targeted analysis to detect the full range of clinically relevant somatic mutations. Here we sought to determine the performance of this targeted analysis approach using long-read sequencing data from Oxford Nanopore Technologies and Pacific Biosciences. Samples from 26 patients with AML and MDS were sequenced to a mean of 52× coverage. Head-to-head comparison of reportable somatic variants to standard WGS revealed more than 96% recall and 91% precision for single nucleotide variants for both long-read platforms. Performance was lower for insertion/deletions (66% recall and 42% precision), especially in regions with few phased reads that facilitate accurate variant detection. The long-read platforms were 95% accurate for copy number calls, and they detected all recurrent structural variants with no false-positive findings. In addition, long reads properly identified intronic insertions near repetitive elements that were incorrectly identified as interchromosomal structural rearrangements by standard WGS. These results indicate that targeted, tumor-only analysis of long-read sequence data is a feasible approach for the genomic evaluation of myeloid cancers, and they show the utility of incorporating variants discovered via long-read sequencing to improve variant interpretation in short-read WGS.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-09-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145187385","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Analytical Validation of a Pan-Cancer Next-Generation Sequencing Assay for In-House Liquid Biopsy Testing: An International Multicenter Study.","authors":"Gaëlle Lescuyer, Alexandre Harlé, Hari Shankar Kumar, Pantelis Constantoulakis, Nicole Pfarr, Ellen Heitzer, Clémence Michon, Gianluca Russo, Ernst-Jan M Speel, Marie Piecyk, Marie Husson, Georgia Christopoulou, Eva-Maria Mayr, Mai-Lan Koppermann, Christophe Passot, Ricarda Graf, Anes Hadjadj Aoul, Violaine Bourdon, Hendrikus J Dubbink, Ronald van Marion, Imke Demers, Anne-Marie C Dingemans, Giancarlo Troncone, Francesco Pepe, Laura Muinelo-Romay, Ángel Díaz-Lagares, Aitor Rodriguez-Casanova, Ramón Manuel Lago Lestón, Deepak Pathak, Parth Shah, Romain V Parillaud, Oskar Martínez de Ilarduya, Jonas Behr, Alexis Rapin, Thomas Vetterli, Sanga Mitra Boppudi, Umberto Malapelle, Léa Payen","doi":"10.1016/j.jmoldx.2025.08.008","DOIUrl":"10.1016/j.jmoldx.2025.08.008","url":null,"abstract":"<p><p>Liquid biopsy assays are transforming precision oncology by providing a less invasive alternative to tissue biopsies. These assays screen for tumor genetic alterations in circulating free DNA, which typically requires detecting variants at low allele frequencies and, therefore, a high sensitivity and specificity. This international, multicenter analytical performance study evaluated the Hedera Profiling 2 circulating tumor DNA test panel (HP2), a hybrid capture-based next-generation sequencing assay for the detection of somatic alterations in circulating free DNA. Covering 32 genes, HP2 enables the detection of single-nucleotide variants (SNVs), insertions and deletions (Indels), fusions, copy number variations, and microsatellite instability status from a single DNA-only workflow. The analytical performance was assessed using reference standards and a diverse cohort of 137 clinical samples precharacterized by orthogonal methods. In reference standards with variants spiked in at 0.5% allele frequency, sensitivity and specificity were 96.92% and 99.67%, respectively, for SNVs/Indels and 100% for fusions. In clinical samples, SNV/Indel detection showed high concordance (94% for European Society for Medical Oncology Scale of Clinical Actionability for Molecular Targets level I variants) with orthogonal methods. Evidence for solid sensitivity was also found for copy number variation detection and microsatellite instability status determination. Overall, the HP2 assay showed significant potential as a sensitive and efficient pan-cancer test for liquid biopsy testing in a decentralized molecular pathology laboratory setting.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145182383","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Validation and Clinical Utility of a Pan-Cancer Circulating Tumor DNA Assay as a First-Approach Test.","authors":"Nisha Kanwar, Michael B Campion, Amber R Schneider, Dragana Milosevic, Carlos Sosa, Antonina A Wojcik, Kevin C Halling, Kandelaria M Rumilla, Ying-Chun Lo, Zhiyv Niu, Katherine B Geiersbach, Margaret A DiGuardo, Benjamin R Kipp, Gang Zheng","doi":"10.1016/j.jmoldx.2025.08.009","DOIUrl":"10.1016/j.jmoldx.2025.08.009","url":null,"abstract":"<p><p>The feasibility of circulating tumor (ct)-DNA assays in first-approach pan-cancer genomic profiling is not well established. Low ctDNA levels limit assay sensitivity, which challenges adaptation to clinical genomic profiling. In this study, a 33-gene next-generation sequencing-based ctDNA panel. The cohorts included 123 patients who underwent first-approach ctDNA testing, and 48 matched patients tested at the same time point. The overall ctDNA assay failure rate was 0%. Insufficient tumor tissue was the main reason for liquid biopsy (69%). The most common primary cancer profiled was lung (39.0%), followed by colon (13.8%), bile duct (8.9%), pancreas (8.1%), and breast and prostate (each 4.1%). Using guidelines from the Association for Molecular Pathology/American Society of Clinical Oncology/College of American Pathologists, Tier I variants were detected in 33.3% of patients, and Tier I or II variants were detected in 65.0% (including 54.5% cholangiocarcinomas, in which tissue biopsy may be challenging due to anatomic location). Compared with matched tissue, ctDNA showed 76% sensitivity for Tier I variants. Actionable variants were increased by 14.3% with ctDNA versus tissue testing alone. ctDNA results preceded tissue results by a mean duration of 21 days. High feasibility, actionability, and sensitivity support ctDNA assays as a potential first-line genomic test, especially in specific tumor types for advanced tumors with insufficient or unavailable tissue.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-09-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145182369","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Next-Generation Sequencing-Based T-Cell Receptor Gene Rearrangement Analysis in Nodal T Follicular Helper Cell Lymphoma, a Comparison with the EuroClonality/BIOMED-2 Assay.","authors":"Ryuko Nakayama, Leonie I Kroeze, Jeroen Luijks, Avital Amir, Jos Rijntjes, Konnie M Hebeda, Patricia J T A Groenen","doi":"10.1016/j.jmoldx.2025.08.007","DOIUrl":"10.1016/j.jmoldx.2025.08.007","url":null,"abstract":"<p><p>Nodal T follicular helper cell lymphoma (nTFHL) can be difficult to diagnose because it often shows features of immune dysregulation and can have a low number of neoplastic T cells in involved lymph nodes. The analysis of T-cell receptor (TR) gene rearrangements by next-generation sequencing (NGS) and the conventional EuroClonality/BIOMED-2 were performed to compare their performance on this challenging diagnosis. DNA was extracted from 32 formalin-fixed, paraffin-embedded nTFHL samples from two pathology archives. NGS amplicon-based analysis of TRBV-TRBD-TRBJ, TRBD-TRBJ, and TRGV-TRGJ rearrangements was performed using the two-step PCR protocols developed by EuroClonality. The nucleotide sequences were analyzed for abundance, clonotype, and functionality. Both the NGS-based and the conventional clonality assays resulted in a high detection of clonality (97% and 94%, respectively), including both monoclonal and biclonal cases. There was an overrepresentation of TRBV20-1 and TRBV19 gene use that was in line with the frequent use of these genes in T cells of reactive lymph nodes and tonsils. The NGS-based approach detected two or more clonal targets in all clonal samples, whereas the conventional assay detected a single (isolated) dominant rearrangement in three cases. Hence, NGS enables more reliable detection of even small clones, as is frequent in nTFHL. NGS-based TR rearrangement analysis provides the abundance, the sequences, clonotypes, and productivity of the clonal rearrangements and thus stresses the need for novel guidelines for interpretation.</p>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":" ","pages":""},"PeriodicalIF":3.4,"publicationDate":"2025-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145151698","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}