Journal of Molecular Diagnostics最新文献

{"title":"Detection of Gene Fusions and Rearrangements in Formalin-Fixed, Paraffin-Embedded Solid Tumor Specimens Using High-Throughput Chromosome Conformation Capture","authors":"Kristyn Galbraith ,&nbsp;Jamin Wu ,&nbsp;Kristin Sikkink ,&nbsp;Hussein Mohamed ,&nbsp;Derek Reid ,&nbsp;Michelle Perez-Arreola ,&nbsp;Jon-Matthew Belton ,&nbsp;Sofia Nomikou ,&nbsp;Shadi Melnyk ,&nbsp;Yiying Yang ,&nbsp;Benjamin L. Liechty ,&nbsp;George Jour ,&nbsp;Aristotelis Tsirigos ,&nbsp;David J. Hermel ,&nbsp;Alyssa Beck ,&nbsp;Darren Sigal ,&nbsp;Nathan A. Dahl ,&nbsp;Rajeev Vibhakar ,&nbsp;Anthony Schmitt ,&nbsp;Matija Snuderl","doi":"10.1016/j.jmoldx.2025.01.007","DOIUrl":"10.1016/j.jmoldx.2025.01.007","url":null,"abstract":"<div><div>Chromosomal structural variants (SVs) are major contributors to cancer development. Although multiple methods exist for detecting SVs, they are limited in throughput, such as fluorescent <em>in situ</em> hybridization and targeted panels, and use RNA, which degrades in formalin-fixed, paraffin-embedded (FFPE) blocks and is unable to detect SVs that do not produce a fusion transcript. High-throughput chromosome conformation capture (Hi-C) is a DNA-based next-generation sequencing (NGS) method that preserves the spatial conformation of the genome, capturing long-range genetic interactions and SVs. Herein, a retrospective study analyzing 71 FFPE specimens from 10 different solid tumors was performed. Results showed high concordance (98%) with clinical fluorescent <em>in situ</em> hybridization and RNA NGS in detecting known SVs. Furthermore, Hi-C provided insight into the mechanism of SV formation, including chromothripsis and extrachromosomal DNA, and detected rearrangements between genes and regulatory regions, all of which are undetectable by RNA NGS. Lastly, SVs were detected in 71% of cases in which previous clinical methods failed to identify a driver. Of these, 14% were clinically actionable based on current medical guidelines, and an additional 14% were not in medical guidelines but involve targetable biomarkers. Current data suggest that Hi-C is a robust and accurate method for genome-wide SV analyses from FFPE tissue and can be incorporated into current clinical NGS workflows.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 5","pages":"Pages 346-359"},"PeriodicalIF":3.4,"publicationDate":"2025-02-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143537013","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 and Comprehensive Whole-Genome Sequencing–Based Preimplantation Genetic Testing Approach for Different Genetic Conditions","authors":"Shuyuan Li ,&nbsp;Chunxin Chang ,&nbsp;Haiyan Bai ,&nbsp;Weiping Qian ,&nbsp;Yangyun Zou ,&nbsp;Dandan Wu ,&nbsp;Wenjing Hu ,&nbsp;Yulin Chen ,&nbsp;Tuan Li ,&nbsp;Sijia Lu ,&nbsp;Wen Li ,&nbsp;Juanzi Shi ,&nbsp;Zhiwei Liu","doi":"10.1016/j.jmoldx.2025.02.002","DOIUrl":"10.1016/j.jmoldx.2025.02.002","url":null,"abstract":"<div><div>Preimplantation genetic testing (PGT) is an essential tool for selecting embryos free of genetic abnormalities. However, current PGT methods often require separate platforms for aneuploidy (PGT-A), monogenic disorders (PGT-M), and structural rearrangements (PGT-SR), leading to increased costs and operational complexity when multiple PGT tests are needed for a single embryo. Here, we present KaryoSeq, a low-pass whole-genome sequencing–based comprehensive PGT approach that integrates PGT-A, PGT-M, and PGT-SR into a single platform. An assistant decision-making system was constructed to pre-evaluate the required sequencing depth for specific genes or regions. Clinical validation of KaryoSeq was performed on 166 blastocyst samples from 31 families previously diagnosed by using conventional PGT methods. KaryoSeq achieved 100% concordance with traditional platforms using the Infinium Asian Screening Array in combination with low-coverage whole-genome sequencing (approximately 0.1×); it also offered improved whole-genome coverage, reduced variability, and efficient simultaneous analysis of PGT-A, PGT-M, and PGT-SR at a whole-genome sequencing depth of approximately 2× for most genes. In addition, KaryoSeq identified triploidy, uniparental disomy, parental origin of copy number variations, and maternal cell contamination, further enhancing its clinical utility and efficiency in PGT applications.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 5","pages":"Pages 395-404"},"PeriodicalIF":3.4,"publicationDate":"2025-02-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143536739","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":"Impact and Reproducibility of In-House Targeted Next-Generation Sequencing Biomarker Testing in Non–Small-Cell Lung Cancer","authors":"Ida Rapa ,&nbsp;Francesca Bertola ,&nbsp;Gaia Roversi ,&nbsp;Davide Seminati ,&nbsp;Federica Panebianco ,&nbsp;Cecília Durães ,&nbsp;Enzo Gallo ,&nbsp;Biagio E. Leone ,&nbsp;Aldo Palange ,&nbsp;Luisella Righi ,&nbsp;Paolo Visca ,&nbsp;Marco Volante ,&nbsp;Simonetta Buglioni","doi":"10.1016/j.jmoldx.2025.02.001","DOIUrl":"10.1016/j.jmoldx.2025.02.001","url":null,"abstract":"<div><div>Next-generation sequencing (NGS) allows the detection of multiple genetic targets in different tumor types. This study aimed to confirm the benefits of implementing in-house NGS testing for non–small-cell lung cancer (NSCLC) samples in molecular pathology laboratories. A multi-institutional study was conducted to evaluate the analytical performance, turnaround time, and feasibility of in-house NGS testing of 50 genes from 283 NSCLC samples. The first phase was a retrospective study with interlaboratory testing (21 samples), and the second phase was a prospective study with intralaboratory testing (262 samples). The retrospective study showed a 100% sequencing success rate for DNA and RNA, high interlaboratory concordance (95.2%), and a strong correlation (<em>R</em>² = 0.94) between observed and expected single-nucleotide variant/insertion and/or deletion variant allele fraction. The prospective study showed a sequencing success rate of 99.2% for DNA and 98% for RNA. NGS identified 285 relevant variants (81.1% single-nucleotide variants/insertion and/or deletion variants, 9.8% copy number variants, and 9.1% gene fusions). Co-mutations with potential clinical relevance were detected in 20.5% of samples positive for the main oncogenic drivers in NSCLC. Additionally, 11% of samples wild type for the main oncogenic drivers carried alterations in other relevant genes. The in-house NGS testing had a median turnaround time from sample processing to molecular report of 4 days. This study demonstrates the advantages of implementing in-house NGS testing in molecular pathology laboratories.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 5","pages":"Pages 371-382"},"PeriodicalIF":3.4,"publicationDate":"2025-02-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143537514","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":"Reviewer Acknowledgment","authors":"","doi":"10.1016/j.jmoldx.2025.01.001","DOIUrl":"10.1016/j.jmoldx.2025.01.001","url":null,"abstract":"","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 3","pages":"Pages 233-234"},"PeriodicalIF":3.4,"publicationDate":"2025-02-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143474168","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":"Retaining Clinical Genomics Technologists in the Post–COVID-19 Era","authors":"Marco L. Leung ,&nbsp;Barbara Anderson","doi":"10.1016/j.jmoldx.2024.10.005","DOIUrl":"10.1016/j.jmoldx.2024.10.005","url":null,"abstract":"","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 3","pages":"Pages 163-165"},"PeriodicalIF":3.4,"publicationDate":"2025-02-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143474288","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":"Repurposing the Whole Expression Transcriptome Assay for the Genetic Diagnosis of T-Cell Acute Lymphoblastic Leukemia and Lymphoma","authors":"Valentina Bardelli ,&nbsp;Silvia Arniani ,&nbsp;Valentina Pierini ,&nbsp;Carlotta Nardelli ,&nbsp;Caterina Matteucci ,&nbsp;Anair Graciela Lema Fernandez ,&nbsp;Maria Crocioni ,&nbsp;Marco Cerrano ,&nbsp;Prassede Salutari ,&nbsp;Cristina Papayanidis ,&nbsp;Silvia Trappolini ,&nbsp;Fabio Giglio ,&nbsp;Sara Mastaglio ,&nbsp;Patrizia Zappasodi ,&nbsp;Crescenza Pasciolla ,&nbsp;Marzia Defina ,&nbsp;Matteo Piccini ,&nbsp;Giuseppe Lanzarone ,&nbsp;Danika Di Giacomo ,&nbsp;Simona Sica ,&nbsp;Roberta La Starza","doi":"10.1016/j.jmoldx.2025.01.006","DOIUrl":"10.1016/j.jmoldx.2025.01.006","url":null,"abstract":"<div><div>Unlike other cases of acute leukemia, the diagnosis of T-cell acute lymphoblastic leukemia/lymphoma (T-ALL/LBL) is uniquely based on morphology and flow cytometry. Although the genomic background has been broadly uncovered, the large spectrum of genes involved and the variability of the molecular mechanisms underlying gene deregulation have delayed the introduction of molecular cytogenetics into diagnostic flowcharts. To overcome these limitations and implement a genetic diagnosis of T-ALL/LBLs, a whole transcriptome expression assay (WTEa) was repurposed as a “priority test” to classify T-ALL/LBLs into the major genetic subtypes. A WTEa classifier based on a set of 312 probes on 215 T-ALL/LBLs was set up and applied, which properly assigned &gt;95% of cases with subtype-defining alterations to the corresponding subgroups (ie, <em>TAL/LMO, HOXA, TLX1, TLX3</em>, <em>BCL11B</em>). It pinpointed cases that harbored cryptic alterations, such as noncoding mutations that generate new enhancer at <em>TAL1</em> and <em>LMO2</em> loci (8% of <em>TAL/LMO</em>), and duplications of noncoding element downstream <em>BCL11B</em> (BETA) (18% of <em>BCL11B</em>). It was also suitable to classify lymphoma cases for which only formalin-fixed embedded tissues were available, as confirmed in cases harboring <em>TLX1</em> or <em>TLX3</em> rearrangements, and distinguished new putative subtypes. WTEa offers a unifying tool to provide a genetic classification of T-ALL/LBLs. If introduced in multicenter prospective studies, it will facilitate evaluation of the clinical impact of genetic classification.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 5","pages":"Pages 360-370"},"PeriodicalIF":3.4,"publicationDate":"2025-02-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143472983","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 Perspective on Artificial Intelligence for Molecular Pathologists","authors":"Timothy J. O'Leary ,&nbsp;Brendan J. O'Leary ,&nbsp;Dianne P. O'Leary","doi":"10.1016/j.jmoldx.2025.01.005","DOIUrl":"10.1016/j.jmoldx.2025.01.005","url":null,"abstract":"<div><div>The widespread adoption of next-generation sequencing technology in molecular pathology has enabled us to interrogate the genome as never before. The huge quantities of data generated by sequencing, the enormous complexity of human and microbial genetics, and the need for fast answers demand increasing use of automation as we diagnose disease and guide patient treatment. Much of this automation is based on tools that fall under umbrellas that have come to be known as machine learning and artificial intelligence. This review outlines some of the broad ideas that underpin these complex computational methods. It discusses the roles of pathologists and data scientists in generating new tools and factors to keep in mind when adopting these systems for use in molecular pathology. It pays special attention to regulatory and professional society guidance for validating them in individual institutions and to possible sources of bias. Finally, it briefly discusses ongoing efforts in computer science that may dramatically impact artificial intelligence in the future.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 5","pages":"Pages 323-335"},"PeriodicalIF":3.4,"publicationDate":"2025-02-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143426564","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":"Biomimetic Digital Twins and Multiomics","authors":"William G. Kearns ,&nbsp;Joe Glick ,&nbsp;Lawrence Baisch ,&nbsp;Andrew Benner ,&nbsp;Dalton Brough ,&nbsp;Luke Du ,&nbsp;Chandra Germain ,&nbsp;Laura Kearns ,&nbsp;Georgios Stamoulis","doi":"10.1016/j.jmoldx.2024.12.012","DOIUrl":"10.1016/j.jmoldx.2024.12.012","url":null,"abstract":"<div><div>The National Academies of Sciences, Engineering, and Medicine issued a report on December 15, 2023, “Foundational Research Gaps and Future Directions for Digital Twins.” This described the importance of using biomimetic digital twins and multiomics in research. These were incorporated in the current analysis of patients with rheumatoid arthritis (RA). Exome sequencing, genotype-phenotype ranking, and biomimetic digital twin analysis were used to identify five pathogenic and one likely pathogenic DNA variants in patient samples analyzed, which were absent from controls. The variants identified in these genes, <em>P2RX7</em>, <em>HTRA2</em>, <em>PTPN22</em>, <em>FLG</em>, <em>CD46</em>, and <em>EIF4G1</em>, play a role in the development of RA. Additionally, 3172 variants of unknown clinical significance (VUSs) were identified in patient samples, which were absent from controls. All VUSs appeared to be associated with RA. Hidden or dark data were identified from six genes. These genes, often found in patient samples, included <em>HIF1A</em>, <em>HLA-DOA</em>, <em>PTGER3</em>, <em>HIPK3</em>, <em>TGFBR3</em>, and <em>HIF1A-AS3</em>. VUSs identified in genes <em>HIF1A</em>, <em>HLA-DOA</em>, <em>PTGER3</em>, and <em>HIPK3</em> were directly related to the pathogenesis of RA, whereas VUSs identified in genes <em>TGFBR3</em> and <em>HIF1A-AS3</em> were indirectly related. The current results suggest that biomimetic digital twins and multiomics can provide further insight into the development of RA. This may also potentially help with the process of reclassifying VUSs. The reclassification of VUSs will play a critical role in complex molecular diagnostics and drug development.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 4","pages":"Pages 256-269"},"PeriodicalIF":3.4,"publicationDate":"2025-02-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143426565","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":"Single-Tube, Switched Temperature Amplicon Barcoding for Multiplex Detection of Rare Mutations in Circulating Tumor DNA","authors":"Tony E. Godfrey ,&nbsp;Ekaterina Kintsurashvili ,&nbsp;Gordana Rasic ,&nbsp;Jessalyn Kaur ,&nbsp;Christopher D'Amato ,&nbsp;Robert H. Meltzer","doi":"10.1016/j.jmoldx.2025.01.004","DOIUrl":"10.1016/j.jmoldx.2025.01.004","url":null,"abstract":"<div><div>Detection and analysis of circulating tumor DNA (ctDNA) as a biomarker for cancer is a promising approach. Applications for ctDNA analysis include screening, diagnosis, treatment selection, treatment monitoring, minimal residual disease detection, and recurrence monitoring. Detection of ctDNA is challenging and requires highly sensitive methods. Approaches such as digital PCR are appropriate when only a small number of targets is being interrogated, whereas next-generation sequencing (NGS) is typically used when more targets are being analyzed. There are several NGS methods available, some of which are published and can be implemented in laboratories with the required expertise while other, commercial approaches are proprietary and are only available as a service. Of the published methods, most use some kind of unique molecular identifiers (or barcodes) to facilitate NGS error correction and detection of rare mutations at mutant allele frequencies of &lt;0.1%. However, incorporation of barcodes and amplification of the resulting libraries are not trivial and typically require multiple steps and considerable hands-on time by an experienced molecular biologist. Herein, a novel approach for switched temperature amplicon barcoding was used, in which barcoding and library amplification were performed in the same tube using a two-stage PCR protocol with no additional manipulation. Total hands-on time was 10 to 15 minutes for reaction setup; the library was then cleaned and was ready for sequencing.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 4","pages":"Pages 237-246"},"PeriodicalIF":3.4,"publicationDate":"2025-02-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143426566","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":"The Validation of Digital PCR–Based Minimal Residual Disease Detection for the Common Mutations in IDH1 and IDH2 Genes in Patients with Acute Myeloid Leukemia","authors":"Jing Di ,&nbsp;Tao Sheng ,&nbsp;Ranjana Arora ,&nbsp;Jennifer Stocks-Candelaria ,&nbsp;Sainan Wei ,&nbsp;Charles Lutz ,&nbsp;Fevzi F. Yalniz ,&nbsp;Shulin Zhang","doi":"10.1016/j.jmoldx.2024.11.002","DOIUrl":"10.1016/j.jmoldx.2024.11.002","url":null,"abstract":"<div><div>Accurate monitoring of minimal residual disease (MRD) is crucial for effective management of patients with acute myeloid leukemia (AML). This study aims to validate MRD detection of the seven most common <em>IDH1</em> and <em>IDH2</em> mutations in patients with AML using a QuantStudio 3D digital PCR platform. This assay demonstrated a high concordance for the variant allele frequencies between digital PCR and next-generation sequencing assays. Precision analysis revealed only small variation (&lt;0.5 log10) for all mutations near or at the limit of detection level. This validation also showed a great reproducibility for interrun and intrarun comparisons (28 runs, variation ranges from 0 to 0.48 log10), ensuring comparable results for patient follow-ups. The limit of detection was determined to be 0.1% for all mutations, except the <em>IDH2 R140Q</em> mutation, which was 0.5%. Controls and acceptable ranges were also established for each mutation during validation. This study suggests that the QuantStudio 3D digital PCR assay is a quantitative, sensitive, and reproducible platform for monitoring MRD in patients with AML.</div></div>","PeriodicalId":50128,"journal":{"name":"Journal of Molecular Diagnostics","volume":"27 2","pages":"Pages 100-108"},"PeriodicalIF":3.4,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142774302","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}