Journal of Molecular Diagnostics最新文献

{"title":"From Expert Knowledge to Validation Resources","authors":"Somak Roy ,&nbsp;Martine W. Tremblay ,&nbsp;Edward Lockhart ,&nbsp;Swaroop Aradhya ,&nbsp;Pinar Bayrak-Toydemir ,&nbsp;Mark Bowser ,&nbsp;Jeana DaRe ,&nbsp;Kristin Gibson ,&nbsp;Michael Kennemer ,&nbsp;Christopher Krueger ,&nbsp;Matt Lebo ,&nbsp;Rong Mao ,&nbsp;Robert Nussbaum ,&nbsp;Brendan O'Fallon ,&nbsp;Andrew Rosato ,&nbsp;Lisa V. Kalman ,&nbsp;Birgit Funke","doi":"10.1016/j.jmoldx.2025.07.006","DOIUrl":"10.1016/j.jmoldx.2025.07.006","url":null,"abstract":"<div><div>Clinical implementation of whole-genome and whole-exome sequencing by next-generation sequencing (NGS) allows for comprehensive detection of genomic alterations. However, with the growing number of clinically relevant genes and variants, there is an urgent need for reference materials to optimize, validate, and quality control NGS tests. This pilot study documents the paucity of physical reference materials for widely tested genes and demonstrates the utility of <em>in silico</em> mutagenized reference materials to supplement physical samples when developing NGS tests. We examined published, expert curated lists of clinically relevant variants for these widely tested genes and found that publicly available reference materials were available for only 29.4%. We outline the steps for generating <em>in silico</em> resources and used 49 curated variants to conduct a blinded proof-of-concept study with three experienced NGS laboratories. One laboratory detected all added variants, and two detected all but one. This study revealed common scenarios that could lead to false-negative results when common pathogenic variants cannot be tested during analytical validation. This work highlights the need to establish centralized knowledge bases for common, pathogenic variants, demonstrates the utility of <em>in silico</em> reference materials, and provides guidance for generating <em>in silico</em> reference materials in-house. Additional work will be needed to generate turnkey processes for novice laboratories without in-house bioinformatics expertise.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 11","pages":"Pages 1074-1083"},"PeriodicalIF":3.4,"publicationDate":"2025-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144976875","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 Atypical Fluorescence in Situ Hybridization Findings by RNA Sequencing","authors":"Beth A. Pitel ,&nbsp;Saba Alvand ,&nbsp;Mark A. Montanari ,&nbsp;Daniel R. Sill ,&nbsp;Carlos Sosa ,&nbsp;Matthew J. Petersen ,&nbsp;Christopher D. Hofich ,&nbsp;Ganesh P. Pujari ,&nbsp;Reid G. Meyer ,&nbsp;Sounak Gupta ,&nbsp;William R. Sukov ,&nbsp;Jorge Torres-Mora ,&nbsp;Kevin C. Halling ,&nbsp;Katherine B. Geiersbach","doi":"10.1016/j.jmoldx.2025.07.008","DOIUrl":"10.1016/j.jmoldx.2025.07.008","url":null,"abstract":"<div><div>Fluorescence <em>in situ</em> hybridization (FISH) using a break-apart probe (BAP) design is a rapid, clinically useful method for targeted evaluation of gene rearrangements in formalin-fixed, paraffin-embedded tumors. Although clinically validated BAP FISH assays usually yield unequivocal positive or negative results, rare tumors yield equivocal FISH results. This study had two aims: to summarize typical and atypical BAP FISH results on 56,584 formalin-fixed, paraffin-embedded solid tumors over approximately one decade of clinical testing; and to investigate the clinical utility of RNA sequencing (RNA-seq) for tumors with equivocal FISH results. Of 8586 (15.2%) cases with abnormal FISH results reported, 748 (8.7%) were equivocal. RNA-seq was performed on 113 tumors, and oncogenic fusions involving the gene of interest were detected in 46 of 113 tumors (40.7%). Of the 106 tumors with equivocal FISH results, RNA-seq detected a fusion involving the expected gene target in 37 of 62 (59.7%) tumors with isolated probe signals corresponding to the active side of the gene region but only 4 of 44 (9.1%) tumors with other atypical signal patterns. This study provides a useful framework for categorizing atypical BAP FISH results and demonstrates the clinical utility of follow-up RNA-seq testing on tumors with equivocal FISH results.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 11","pages":"Pages 1098-1114"},"PeriodicalIF":3.4,"publicationDate":"2025-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144994166","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 Genomic Proximity Mapping for Detecting Genomic and Chromosomal Structural Variants in Constitutional Disorders","authors":"He Fang ,&nbsp;Stephen M. Eacker ,&nbsp;Yu Wu ,&nbsp;Cate Paschal ,&nbsp;Mary Wood ,&nbsp;Brad Nelson ,&nbsp;Alexander Muratov ,&nbsp;Yajuan J. Liu","doi":"10.1016/j.jmoldx.2025.07.005","DOIUrl":"10.1016/j.jmoldx.2025.07.005","url":null,"abstract":"<div><div>Structural variants are critical to genetic diversity and disease, yet their detection remains challenging with conventional cytogenetic techniques, including karyotyping, fluorescence <em>in situ</em> hybridization, and chromosome microarray analysis. These methods often lack the resolution and sensitivity needed for comprehensive characterization of chromosomal aberrations. To address these limitations, genomic proximity mapping (GPM), a genome-wide chromosome conformation capture (Hi-C) technology, was implemented in a clinical setting. Here, GPM was applied to a cohort of 123 individuals with constitutional disorders, achieving a 100% concordance in detecting 110 copy number variants (&gt;25 kb) and 27 structural rearrangements, in addition to novel findings. GPM demonstrated unique advantages, such as resolving chromosomal rearrangements with precise (&lt;5 kb) breakpoint resolution, maintaining robust performance with challenging samples, including formalin-fixed, paraffin-embedded tissues, and detecting mosaicism with high sensitivity. Furthermore, GPM reliably provided copy number and loss-of-heterozygosity profiles, streamlining workflows. GPM also uncovered 12 novel structural variants missed by standard methods, highlighting its superior detection capability. This analysis revealed that cases with more than two chromosomal rearrangements identified by traditional cytogenetics often harbor additional, cryptic rearrangements that remain undetected by standard-of-care methods. GPM represents a transformative tool for genomic diagnostics, offering a high-resolution, comprehensive approach to detecting genomic alterations. Its ability to address limitations of conventional cytogenetics methods positions GPM as a needed advance in the diagnosis, prognosis, and therapeutic management of genetic disorders.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 11","pages":"Pages 1054-1069"},"PeriodicalIF":3.4,"publicationDate":"2025-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144976902","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":"Validating Fluorescence in Situ Hybridization with RNA Sequencing","authors":"Fei Dong","doi":"10.1016/j.jmoldx.2025.08.004","DOIUrl":"10.1016/j.jmoldx.2025.08.004","url":null,"abstract":"","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 11","pages":"Pages 1037-1038"},"PeriodicalIF":3.4,"publicationDate":"2025-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145006752","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":"FoundationOne CDx and FoundationOne Heme Detect Epstein-Barr Virus with High Sensitivity and Specificity","authors":"Julius Honecker ,&nbsp;Dirk Paffenholz ,&nbsp;Julian Jungmann ,&nbsp;Thomas Wieland ,&nbsp;Angela A. Kou ,&nbsp;Soo-Ryum Yang ,&nbsp;Chad M. Vanderbilt ,&nbsp;Vignesh Shanmugam ,&nbsp;Meagan Montesion ,&nbsp;Ryon P. Graf ,&nbsp;Lee A. Albacker ,&nbsp;Douglas I. Lin ,&nbsp;Andreas Heilmann ,&nbsp;Sotirios Lakis ,&nbsp;Richard A. Hickman ,&nbsp;Brennan Decker ,&nbsp;Alex Robertson ,&nbsp;Lauren L. Ritterhouse ,&nbsp;Douglas A. Mata","doi":"10.1016/j.jmoldx.2025.06.002","DOIUrl":"10.1016/j.jmoldx.2025.06.002","url":null,"abstract":"<div><div>Accurate detection of Epstein-Barr virus (EBV) in tumors is essential for refining diagnosis and guiding management of EBV-associated cancers. This validation study evaluated the performance of the FoundationOne CDx (F1CDx) and FoundationOne Heme (F1H) next-generation sequencing assays for detecting EBV compared with the reference standard EBV-encoded RNA (EBER) <em>in situ</em> hybridization (ISH). A total of 413 samples of gastric carcinoma, squamous cell carcinoma (predominantly nasopharyngeal carcinoma), and diffuse large B-cell lymphoma was assessed. The assays demonstrated a sensitivity of 94.0%, specificity of 99.1%, and accuracy of 98.3% with an area under the curve of 0.97 for EBV detection, showing high concordance with EBER ISH. Precision was 95.5%, and reproducibility across multiple runs was strong (CV = 4.7%). Dilution experiments demonstrated a linear relationship between EBV viral reads per million and the degree of cell culture dilution (Pearson's R &gt; 0.99, <em>P</em> &lt; 0.001), supporting assay linearity over a broad dynamic range. Characteristic mutational and cytogenetic profiles, including losses at 9p21 and gains at 11q13, were observed in EBV-positive samples, supporting the clinical utility of the method. These results indicate that F1CDx and F1H provide a reliable, in-parallel approach for detecting EBV along with somatic genomic alterations in a single assay. This method offers a promising alternative to conventional ISH, especially for identifying EBV in tumors without initial clinical suspicion, potentially streamlining diagnostic workflows.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 10","pages":"Pages 920-935"},"PeriodicalIF":3.4,"publicationDate":"2025-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144719035","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 Implementation of Matched Tumor/Germline Sequencing Improves Accuracy of Tumor Genomic Profiling and Therapeutic Recommendations","authors":"Danielle K. Manning ,&nbsp;Guruprasad Ananda ,&nbsp;Cheryl Eifert ,&nbsp;Vanesa Rojas-Rudilla ,&nbsp;William Swanton ,&nbsp;Shawn Keefe ,&nbsp;Elizabeth P. Garcia ,&nbsp;Michael D'Eletto ,&nbsp;Erica Holdmore ,&nbsp;Satyakam Mishra ,&nbsp;Kirill Borziak ,&nbsp;Pieter Lukasse ,&nbsp;Phani Davineni ,&nbsp;Monica D. Manam ,&nbsp;Priyanka Shivdasani ,&nbsp;Arezou A. Ghazani ,&nbsp;Ai Ling Wang ,&nbsp;Murat Bastepe ,&nbsp;Himisha Beltran ,&nbsp;Katherine A. Janeway ,&nbsp;Laura E. MacConaill","doi":"10.1016/j.jmoldx.2025.06.005","DOIUrl":"10.1016/j.jmoldx.2025.06.005","url":null,"abstract":"<div><div>Genomic profiling of cancers informs diagnostic and prognostic classification and aids in selection of targeted therapeutics. Targeted, next-generation sequencing of cancer-specific genes is clinically feasible and enables comprehensive somatic reporting; without a matched germline specimen, germline alterations can confound analyses of the somatic profile and generate uncertainty in interpretation. This work reports the validation and implementation of optional matched tumor/germline sequencing in a precision cancer medicine program. DNA from 63 patient samples was analyzed using OncoPanel, a hybrid capture-based sequencing assay of 461 genes. Three analytical pipelines were implemented: tumor only, matched tumor/germline, and germline only. For matched tumor/germline, germline alterations in 19 genes with actionable/therapeutic implications were rescued. Retrospective analysis of the first 1600 matched cases was done to determine the potential clinical utility of this approach. Limit of detection for point mutations/insertions and deletions was 3% allele fraction; reproducibility was &gt;98%. Matched tumor/germline concordance across 938 somatic calls was 100%. The average tumor mutational burden (TMB) was approximately 4 mutations/Mb lower than tumor-only sequencing. TMB-high patients were accurately reclassified as TMB-low in 14% of cases. Twenty-five percent of validation cases (14% after launch) had a pathogenic or likely pathogenic germline variant conferring cancer susceptibility; 14% of validation cases (7% after launch) harbored a germline variant of therapeutic significance. Matched tumor/germline sequencing is more accurate than tumor-only sequencing, while still encompassing all genomic findings that inform targeted therapy selection.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 10","pages":"Pages 969-988"},"PeriodicalIF":3.4,"publicationDate":"2025-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144709677","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":"Bridging the Diagnostic Gap in Hereditary Cancers with Simple, Cost-Effective, High-Throughput RNA Splicing Analysis","authors":"Julie Amiot ,&nbsp;Corentin Levacher ,&nbsp;Louise May Thibaut ,&nbsp;Gwendoline Lienard ,&nbsp;Stéphanie Vasseur ,&nbsp;Olivier Quenez ,&nbsp;Sophie Coutant ,&nbsp;Steeve Fourneaux ,&nbsp;Françoise Charbonnier ,&nbsp;Hugo Thorn ,&nbsp;Angélina Legros ,&nbsp;Camille Aucouturier ,&nbsp;Laurent Castéra ,&nbsp;Raphaël Leman ,&nbsp;Edwige Kasper ,&nbsp;Stéphanie Baert-Desurmont ,&nbsp;Sophie Krieger ,&nbsp;Philippe Ruminy ,&nbsp;Claude Houdayer","doi":"10.1016/j.jmoldx.2025.06.003","DOIUrl":"10.1016/j.jmoldx.2025.06.003","url":null,"abstract":"<div><div>Diagnostic analysis of mRNA is essential because altered splicing is a frequent cause of genetic diseases. High-throughput splicing studies remain difficult to implement in routine diagnostics. This is why SEALigHTS (splice and expression analyses by ligation and high throughput sequencing), a cost-effective and easy-to-implement technique designed for simultaneous analysis of RNA from multiple patients on a panel of genes, was developed using probes designed at exon extremities. After reverse transcription and probing on cDNA, neighboring probes are ligated and the number of ligations quantified by using unique molecular identifiers and sequencing. A panel covering 42 genes (ie, 2195 probes) involved in breast/ovarian and colorectal cancer predispositions was designed. After a training phase on 40 samples, SEALigHTS was validated in another laboratory on 56 samples carrying various splicing variations previously characterized by RNA sequencing, with a sensitivity of 96% and specificity of 94%. Subsequently, in a series of 37 selected patients and 114 consecutive patients from the genetics clinic with a concomitant DNA panel, five new diagnoses were made, revealing the impact on splicing of a cryptic genomic variant (deep intronic, retrotransposon insertion), and the unexpected impact on splicing of six genomic variations was unmasked. Beyond increased diagnostic yield and classification of variants of uncertain significance, this comprehensive DNA and RNA combined approach highlights unexpected splicing defects and addresses genotype–phenotype correlation issues.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 10","pages":"Pages 954-968"},"PeriodicalIF":3.4,"publicationDate":"2025-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144719034","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":"Celebrating 25 Years of Innovation with a Tribute to Our Contributors and the Future of Molecular Diagnostics","authors":"Ronald M. Przygodzki (Editor-in-Chief)","doi":"10.1016/j.jmoldx.2025.08.001","DOIUrl":"10.1016/j.jmoldx.2025.08.001","url":null,"abstract":"","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 10","pages":"Page 913"},"PeriodicalIF":3.4,"publicationDate":"2025-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145121100","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 Blood-Derived Tumor Mutation Burden (bTMB) Assays","authors":"Jonathan Baden ,&nbsp;Mark Sausen ,&nbsp;Andrew T. Anfora ,&nbsp;Kevin M. D'Auria ,&nbsp;Jennifer Dickey ,&nbsp;James H. Godsey ,&nbsp;Li Guan ,&nbsp;Jennifer S. Lococo ,&nbsp;Elizabeth Mansfield ,&nbsp;Kristen L. Meier ,&nbsp;David Merriam ,&nbsp;Traci Pawloski ,&nbsp;Soni Shukla ,&nbsp;Daniel Stetson ,&nbsp;Mark D. Stewart ,&nbsp;Paul Wenz ,&nbsp;Lauren C. Leiman","doi":"10.1016/j.jmoldx.2025.06.008","DOIUrl":"10.1016/j.jmoldx.2025.06.008","url":null,"abstract":"<div><div>Immunotherapies have changed the treatment paradigm for patients with advanced and metastatic solid tumors, with tumor mutation burden representing one approach to identify patients who may experience clinical benefit. Circulating tumor DNA–based approaches have been developed for comprehensive analyses of clinically actionable biomarkers; however, blood tumor mutation burden (bTMB) represents a novel, complex biomarker. Although the clinical utility of bTMB is an evolving area of active development and has not led to consistent conclusions across studies, robust analytical validation of the underlying test is important to ensure that technical and biological limitations do not confound clinical interpretation of these results. To this end, the BLOODPAC bTMB Analytical Validation Working Group sought to identify key technical and biological issues associated with analytical validation of bTMB tests, along with a conceptual framework to address these challenges. This publication provides guidance for device manufacturers to demonstrate analytical performance of their test with the understanding that these data would be accompanied by an appropriately designed clinical validation study to demonstrate performance within the intended use population. Therefore, the specific algorithm to determine the bTMB result, along with the associated cutoff, is out of scope of this Perspective. Device manufacturers should also ensure that appropriate pre-analytical variables are accounted for and methods are incorporated to differentiate tumor-specific alterations from those associated with germline polymorphisms or clonal hematopoiesis.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 10","pages":"Pages 914-919"},"PeriodicalIF":3.4,"publicationDate":"2025-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145121101","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":"A Novel Droplet Digital PCR Assay for BRCA1 and RAD51C Methylation","authors":"Cassandra Michel ,&nbsp;Alizée Simon ,&nbsp;Andréa Witz ,&nbsp;Idrissia Hanriot ,&nbsp;Julie Dardare ,&nbsp;Margaux Betz ,&nbsp;Maxime Brun ,&nbsp;Pauline Gilson ,&nbsp;Jean Louis Merlin ,&nbsp;Alexandre Harlé","doi":"10.1016/j.jmoldx.2025.07.002","DOIUrl":"10.1016/j.jmoldx.2025.07.002","url":null,"abstract":"<div><div>Homologous recombination deficiency (HRD) affects 50% of ovarian cancers and influences poly (ADP-ribose) polymerase inhibitor efficacy. Although one-third of HRD tumors harbor a deleterious <em>BRCA1/2</em> mutation, these mutations are not the sole cause of HRD. Promoter methylation of <em>BRCA1</em> and <em>RAD51C</em> contributes to 19% and 2% of cases, respectively. A cohort of 224 patients with ovarian cancer tested for HRD validated the droplet digital PCR (ddPCR) technique. DNA was extracted from formalin-fixed, paraffin-embedded tissue, and the Genomic Instability Index (GII) was assessed using the SOPHiA DDM HRD Solution. <em>BRCA1</em> and <em>RAD51C</em> promoter methylation was analyzed by ddPCR following enzymatic conversion, with a 10% threshold for methylation classification. Homologous recombination status was assessed in 194 patients. A positive GII (&gt;0) was identified in 77 cases (39.7%), with 24 (31.2%) harboring a deleterious <em>BRCA1</em>/2 mutation, and 2 (2.6%) a <em>RAD51C</em> mutation. Methylation analysis revealed <em>BRCA1</em> promoter methylation in 32 patients (41.6%) and <em>RAD51C</em> promoter methylation in 1 patient (1.3%), clarifying HRD status in 28 additional cases (36.3%). The study confirms HRD extends beyond <em>BRCA1</em>/2 mutations, with promoter methylation playing a significant role in HRD detection. ddPCR effectively identified methylation-driven HRD, explaining deficiency in 36.4% additional patients with a positive GII score. These findings highlight the importance of incorporating methylation analysis into HRD testing to improve patient stratification for poly (ADP-ribose) polymerase inhibitor therapy.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 10","pages":"Pages 1026-1036"},"PeriodicalIF":3.4,"publicationDate":"2025-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144812620","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}