Preanalytical Variables for the Genomic Assessment of the Cellular and Acellular Fractions of the Liquid Biopsy in a Cohort of Breast Cancer Patients
Liquid biopsy allows assessment of multiple analytes, providing temporal information with potential for improving understanding of cancer evolution and clinical management of patients. Although liquid biopsies are intensely investigated for prediction and response monitoring, preanalytic variables are of primary concern for clinical implementation, including categories of collection method and sample storage. Herein, an integrated high-density single-cell assay workflow for morphometric and genomic analysis of the liquid biopsy is used to characterize the effects of preanalytical variation and reproducibility of data from ...
Source: Journal of Molecular Diagnostics - January 21, 2020 Category: Pathology Authors: Stephanie N. Shishido, Lisa Welter, Mariam Rodriguez-Lee, Anand Kolatkar, Liya Xu, Carmen Ruiz, Anna S. Gerdtsson, Sara Restrepo-Vassalli, Anders Carlsson, Joe Larsen, Emily J. Greenspan, E. Shelley Hwang, Kathryn R. Waitman, Jorge Nieva, Kelly Bethel, Ja Tags: Regular article Source Type: research

An Isothermal, Multiplex Amplification Assay for Detection and Genotyping of Human Papillomaviruses in Formalin-Fixed, Paraffin-Embedded Tissues
Rapid and accurate identification of human papillomavirus (HPV) is important for both clinical management and population screening. We performed analytic validation of Atila AmpFire Multiplex HPV assays on formalin-fixed, paraffin-embedded (FFPE) cervix/vulva and oropharynx diagnostic tissue samples. The AmpFire assay incorporates a novel isothermal multiplex amplification coupled with real-time fluorescent detection to detect and genotype 15 high-risk (HR) HPV genotypes. Limits of detection determined by plasmids cloned with HPV genotype-specific sequences were 2 copies/reaction for HPV16, HPV18, and some HR HPV genotypes...
Source: Journal of Molecular Diagnostics - January 21, 2020 Category: Pathology Authors: Yi-Wei Tang, Lorena Lozano, Xin Chen, Troy D. Querec, Nora Katabi, Antonio Moreno-Doc ón, Hongmei Wang, Daniel Fix, Louise De Brot, Tracy A. McMillen, Ju-Yoon Yoon, Amparo Torroba, Youxiang Wang, Elizabeth R. Unger, Kay J. Park Tags: Regular Article Source Type: research

Diagnosing Bacteremia in Real Time Using Next-Generation Sequencing –Based Technology
This commentary highlights the article by Grumaz et al that describes the use of molecular sequencing for fast detection of pathogens directly from blood samples from septic patients. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - January 21, 2020 Category: Pathology Authors: Charles W. Stratton, Yi-Wei Tang Tags: Commentary Source Type: research

Pre-analytical variables for the genomic assessment of the cellular and acellular fractions of the liquid biopsy in a cohort of breast cancer patients
The liquid biopsy allows assessment of multiple analytes providing temporal information with potential for improving our understanding of cancer evolution and clinical management of patients. While liquid biopsies are intensely investigated for prediction and response monitoring, pre-analytic variables are of primary concern for clinical implementation, including categories of collection methodology and sample storage. Here we utilize an integrated high-definition single cell assay (HD-SCA) workflow for morphometric and genomic analysis of the liquid biopsy to characterize the effects of pre-analytical variation and reprod...
Source: Journal of Molecular Diagnostics - January 21, 2020 Category: Pathology Authors: Stephanie N. Shishido, Lisa Welter, Mariam Rodriguez-Lee, Anand Kolatkar, Liya Xu, Carmen Ruiz, Anna S. Gerdtsson, Sara Restrepo-Vassalli, Anders Carlsson, Joe Larsen, Emily J. Greenspan, E. Shelley Hwang, Kathryn R. Waitman, Jorge Nieva, Kelly Bethel, Ja Tags: Regular Article Source Type: research

Low Pass Whole Genome Sequencing as a Method of Determining Copy Number Variations in Uveal Melanoma Tissue Samples
Analysis of specific somatic copy number alterations (SCNAs) using multiplex ligation-dependent probe amplification (MLPA) is routinely used as prognostic test for uveal melanoma (UM). This technique requires relatively large amounts of input DNA, unattainable from many small fine needle aspirate biopsies. Herein we compared the use of MLPA with whole genome amplification (WGA) combined with low-pass whole genome sequencing (LP-WGS) for detection of SCNA profiles in UM biopsy specimens. DNA was extracted from 21 formalin-fixed paraffin-embedded FFPE UM samples and SCNAs were assessed using MLPA and WGA followed by LP-WGS. ...
Source: Journal of Molecular Diagnostics - January 21, 2020 Category: Pathology Authors: Aaron B. Beasley, Jacqueline Bentel, Richard J.N. Allcock, Tersia Vermeulen, Leslie Calapre, Timothy Isaacs, Melanie R. Ziman, Fred K. Chen, Elin S. Gray Tags: Regular Article Source Type: research

Junction Location Identifier (JuLI)
Accurate detection of genomic fusions by high-throughput sequencing in clinical samples with inadequate tumor purity and formalin-fixed, paraffin-embedded tissue is an essential task in precise oncology. We developed the fusion detection algorithm Junction Location Identifier (JuLI) for optimization of high-depth clinical sequencing. Novel filtering steps were implemented to minimize false positives in the clinical setting. The algorithm was comprehensively validated using high-depth sequencing data from cancer cell lines and clinical samples and genome sequencing data from NA12878. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 24, 2019 Category: Pathology Authors: Hyun-Tae Shin, Nayoung K.D. Kim, Jae Won Yun, Boram Lee, Sungkyu Kyung, Ki-Wook Lee, Daeun Ryu, Jinho Kim, Joon Seol Bae, Donghyun Park, Yoon-La Choi, Se-Hoon Lee, Myung-Ju Ahn, Keunchil Park, Woong-Yang Park Tags: Regular article Source Type: research

Validation of an in  Vitro Mismatch Repair Assay Used in the Functional Characterization of Mismatch Repair Variants
A significant proportion of DNA-mismatch repair (MMR) variants are classified as of unknown significance, precluding diagnosis. The in  vitro MMR assay is used to assess their MMR capability, likely the most important function of an MMR protein. However, the robustness of the assay, crucial for its use in the clinical setting, has been rarely evaluated. The aim of the present work was to validate an in vitro MMR assay approach to the functional characterization of MMR variants, as a first step to meeting quality standards of diagnostic laboratories. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 24, 2019 Category: Pathology Authors: Maribel Gonz ález-Acosta, Inga Hinrichsen, Anna Fernández, Conxi Lázaro, Marta Pineda, Guido Plotz, Gabriel Capellá Tags: Regular article Source Type: research

Junction Location Identifier
Accurate detection of genomic fusions by high-throughput sequencing in clinical samples with inadequate tumor purity and formalin-fixed, paraffin-embedded tissue is an essential task in precise oncology. We developed the fusion detection algorithm Junction Location Identifier for optimization of high-depth clinical sequencing. Novel filtering steps were implemented to minimize false positives in the clinical setting. The algorithm was comprehensively validated using high-depth sequencing data from cancer cell lines and clinical samples and genome sequencing data from NA12878. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 24, 2019 Category: Pathology Authors: Hyun-Tae Shin, Nayoung K.D. Kim, Jae Won Yun, Boram Lee, Sungkyu Kyung, Ki-Wook Lee, Daeun Ryu, Jinho Kim, Joon Seol Bae, Donghyun Park, Yoon-La Choi, Se-Hoon Lee, Myung-Ju Ahn, Keunchil Park, Woong-Yang Park Tags: Regular article Source Type: research

Apparently Heterozygous TP53 Pathogenic Variants May Be Blood Limited in Patients Undergoing Hereditary Cancer Panel Testing
Heterozygous (HET) TP53 pathogenic variants (PVs) are associated with Li-Fraumeni syndrome (LFS), a dominantly inherited condition causing high risk for sarcoma, breast, and other cancers. Recent reports described patients without features of LFS and apparently HET TP53 PVs in blood cells but not fibroblasts (FBs), suggesting the variant occurred sporadically during hematopoiesis and rose to high allele fraction through clonal expansion. To explore possible clonal hematopoiesis in patients undergoing hereditary cancer testing, FB testing was performed for patients with apparently HET or mosaic TP53 PVs identified in blood,...
Source: Journal of Molecular Diagnostics - December 24, 2019 Category: Pathology Authors: Jessica L. Mester, Sarah A. Jackson, Kristen Postula, Amy Stettner, Sheila Solomon, Jeffrey Bissonnette, Patricia D. Murphy, Rachel T. Klein, Kathleen S. Hruska Tags: Regular article Source Type: research

Reliable Clinical MLH1 Promoter Hypermethylation Assessment using a High-Throughput Genome-Wide Methylation Array Platform
Clinical testing for MLH1 promoter hypermethylation status is important in the workup of patients with MLH1-deficient colorectal and uterine carcinomas when evaluating patients for Lynch Syndrome. Current assays utilize single gene –based methods to assess promoter hypermethylation. Herein, we describe the development, and report the performance of a clinical assay for MLH1 promoter hypermethylation using the Infinium methylationEPIC (850k) bead-array platform. Using four CpG sites within the MLH1 gene promoter, a qualitativ e MLH1 promoter hypermethylation assay was developed and validated using 63 gastrointestinal ...
Source: Journal of Molecular Diagnostics - December 24, 2019 Category: Pathology Authors: Jamal K. Benhamida, Jaclyn F. Hechtman, Khedoudja Nafa, Liliana Villafania, Justyna Sadowska, Jiajing Wang, Donna Wong, Ahmet Zehir, Liying Zhang, Tejus Bale, Maria E. Arcila, Marc Ladanyi Source Type: research

JuLI: accurate detection of DNA fusions in clinical sequencing for precision oncology
Accurate detection of genomic fusions by high-throughput sequencing in clinical samples with inadequate tumor purity and formalin-fixed, paraffin embedded tissue is an essential task in precise oncology. We developed the fusion detection algorithm Junction Location Identifier for optimization of high-depth clinical sequencing. Novel filtering steps were implemented to minimize false positives in clinical setting. The algorithm was comprehensively validated using high-depth sequencing data from cancer cell lines and clinical samples and genome sequencing data from NA12878. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 24, 2019 Category: Pathology Authors: Hyun-Tae Shin, Nayoung K.D. Kim, Jae Won Yun, Boram Lee, Sungkyu Kyung, Ki-Wook Lee, Daeun Ryu, Jinho Kim, Joon Seol Bae, Donghyun Park, Yoon-La Choi, Se-Hoon Lee, Myung-Ju Ahn, Keunchil Park, Woong-Yang Park Tags: Regular Article Source Type: research

Validation of an in vitro mismatch repair assay used in the functional characterization of mismatch repair variants
A significant proportion of DNA mismatch repair (MMR) variants are classified as of unknown significance (VUS), precluding diagnosis. The in vitro MMR assay is used to assess their MMR capability, likely the most important function of a MMR protein. However, robustness of the assay, critical for its use in the clinical setting, has been rarely evaluated. The aim of the present work was to validate an in vitro MMR assay approach for the functional characterization of MMR variants, as a first step to meeting quality standards of diagnostic laboratories. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 24, 2019 Category: Pathology Authors: Maribel Gonz ález-Acosta, Inga Hinrichsen, Anna Fernández, Conxi Lázaro, Marta Pineda, Guido Plotz, Gabriel Capellá Tags: Regular Article Source Type: research

Clinical performance of the Idylla MSI Test for a rapid assessment of the DNA microsatellite status in human colorectal cancer
In this study we evaluated the clinical performance of the Idylla MSI Test (Investigational Use Only) in 330 colorectal carcinoma samples (all stages). This test is fully automated from formalin-fixed, paraffin-embedded slide to result, and gives a result in less than 2.5 hours. Compared to the Promega MSI Analysis System, Version 1.2, an overall agreement, sensitivity, and specificity of 99.7%, 98.7%, and 100% was reached, respectively. Whereas seven samples were invalid with the Promega MSI Analysis, only two were invalid with the Idylla MSI Test. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 24, 2019 Category: Pathology Authors: Karen Zwaenepoel, Julie Holmgaard Duelund, Koen De Winne, Vincent Maes, Christine Weyn, Suzan Lambin, Robina Dendooven, Glenn Broeckx, Torben Steiniche, Patrick Pauwels Tags: Regular Article Source Type: research

Apparently Heterozygous TP53 Pathogenic Variants May Be Blood-Limited in Patients Undergoing Hereditary Cancer Panel Testing
Heterozygous (HET) TP53 pathogenic variants (PV) are associated with Li-Fraumeni syndrome (LFS), a dominantly inherited condition causing high risk for sarcoma, breast, and other cancers. Recent reports have described patients without features of LFS and apparently HET TP53 PV in blood cells but not fibroblasts (FB), suggesting the variant occurred sporadically during hematopoiesis and rose to high allele fraction through clonal expansion. To explore possible clonal hematopoiesis in patients undergoing hereditary cancer testing, FB testing was performed for patients with apparently HET or mosaic TP53 PV identified in blood...
Source: Journal of Molecular Diagnostics - December 24, 2019 Category: Pathology Authors: Jessica L. Mester, Sarah A. Jackson, Kristen Postula, Amy Stettner, Sheila Solomon, Jeffrey Bissonnette, Patricia D. Murphy, Rachel T. Klein, Kathleen S. Hruska Tags: Regular Article Source Type: research

Validation of Fragile X Screening in the Newborn Population Using a Fit-for-Purpose FMR1 PCR Assay System
Newborn screening is designed for pre-symptomatic identification of serious conditions with effective early interventions. Clinical laboratories must perform prospective pilot studies to ensure that the analytical performance and workflow for a given screening test are appropriate. We assessed the potential to screen newborns for fragile X syndrome, a monogenic neurodevelopmental disorder, by establishing a customized, high-throughput PCR and analysis software system designed to detect fragile X mental retardation 1 gene repeat expansions from dried blood spots. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 19, 2019 Category: Pathology Authors: Stacey Lee, Jennifer L. Taylor, Charles Redmond, Andrew Hadd, Jon A. Kemppainen, Brian C. Haynes, Scott Shone, Donald B. Bailey, Gary J. Latham Tags: Regular Article Source Type: research

Development and Analytical Validation of an Expanded Mutation Detection Panel for Next-Generation Sequencing of Thyroid Nodule Aspirates
Molecular analysis is used to evaluate the risk of malignancy for thyroid fine needle aspirates, identified as indeterminate by microscopic cytology, based on the detection of various oncogenic DNA mutations and fusion transcripts, or upon the use of various mRNAs or microRNA-based classifier algorithms. Our approach has been to use a combination test utilizing the detection of oncogenic mutations/ fusion transcripts and a micro-RNA expression –based classifier algorithm. To improve the performance of the combination test, the next-generation sequencing (NGS)-based mutational panel was expanded from the detection of ...
Source: Journal of Molecular Diagnostics - December 19, 2019 Category: Pathology Authors: Kenny Kwabena Ablordeppey, Venkata Arun Timmaraju, Joanna Wanmin Song-Yang, Sharon Yaqoob, Christina Narick, Alidad Mireskandari, Sydney David Finkelstein, Gyanendra Kumar Tags: Regular article Source Type: research

Molecular discordance between myeloid sarcomas and concurrent bone marrows occurs in actionable genes and is associated with worse overall survival
Myeloid sarcoma is a rare, architecture-effacing proliferation of myeloid blasts localized to an extramedullary site, with or without concurrent bone marrow involvement. Clonal heterogeneity results from acquisition of somatic mutations within different subclones of leukemic cells. We hypothesized that clonal heterogeneity between myeloid sarcomas and concurrent bone marrow biopsies might be present given their differing biologic features and microenvironment. High-throughput sequencing of the largest series (n = 24) of paired myeloid sarcomas and bone marrow biopsies was performed. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 19, 2019 Category: Pathology Authors: Brian Werstein, Jennifer Dunlap, Michael J. Cascio, Robert S. Ohgami, Guang Fan, Richard Press, Philipp W. Raess Source Type: research

Renewed Commitments to Key Partners
This Editorial highlights the updates to the publication frequency, member benefits, and societal oversight of The Journal of Molecular Diagnostics beginning in January  2020. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 18, 2019 Category: Pathology Authors: Barbara A. Zehnbauer Tags: Editorial Source Type: research

Editorial Board
(Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 18, 2019 Category: Pathology Source Type: research

Table of Contents
(Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 18, 2019 Category: Pathology Source Type: research

Instructions to Authors
(Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 18, 2019 Category: Pathology Source Type: research

Scientific Integrity Policy
(Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 18, 2019 Category: Pathology Source Type: research

Acoustofluidic Salivary Exosome Isolation
In this study, an acoustofluidic (the fusion of acoustics and microfluidics) platform was developed to perform size-based isolation of salivary exosomes. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 13, 2019 Category: Pathology Authors: Zeyu Wang, Feng Li, Joseph Rufo, Chuyi Chen, Shujie Yang, Liang Li, Jinxin Zhang, Jordan Cheng, Yong Kim, Mengxi Wu, Elliot Abemayor, Michael Tu, David Chia, Rachel Spruce, Nikolaos Batis, Hisham Mehanna, David T.W. Wong, Tony Jun Huang Tags: Regular article Source Type: research

Concordance of Genomic Alterations by Next-Generation Sequencing in Tumor Tissue versus Cell-Free DNA in Stage I –IV Non–Small Cell Lung Cancer
Molecular biomarkers hold promise for personalization of cancer treatment. However, a typical tumor biopsy may be difficult to acquire and may not capture genetic variations within or across tumors. Given these limitations, tumor genotyping using next-generation sequencing of plasma-derived circulating tumor (ct)-DNA has the potential to transform non –small cell lung cancer (NSCLC) management. Importantly, mutations detected in biopsied tissue must also be detected in plasma-derived ctDNA at different disease stages. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: John Jiang, Hans-Peter Adams, Lijing Yao, Stephanie Yaung, Preeti Lal, Aarthi Balasubramanyam, Frederike Fuhlbr ück, Nalin Tikoo, Alexander F. Lovejoy, Sebastian Froehler, Li Tai Fang, H. Jost Achenbach, Ralph Floegel, Rainer Krügel, John Palma Tags: Regular article Source Type: research

Development and Validation of a Novel and Rapid Molecular Detection Method for High-Risk Human Papillomavirus in Formalin-Fixed Paraffin-Embedded Tumor Tissue
In this study, we developed and validated a molecular HPV detection method for FFPE specimens of oropharyngeal and nonoropharyngeal HNSCC. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Steven W. Mes, Dani ëlle A.M. Heideman, Elisabeth Bloemena, Arjen Brink, Martijn Bogaarts, C. René Leemans, Ruud H. Brakenhoff Tags: Regular article Source Type: research

Development and Analytical Validation of a DNA Dual-Strand Approach for the US Food and Drug Administration –Approved Next-Generation Sequencing–Based Praxis Extended RAS Panel for Metastatic Colorectal Cancer Samples
A next-generation sequencing method was developed that can distinguish single-stranded modifications from low-frequency somatic mutations present on both strands of DNA in formalin-fixed paraffin-embedded colorectal cancer samples. We applied this method for analytical validation of the Praxis Extended RAS Panel, a US Food and Drug Administration –approved companion diagnostic for panitumumab, on the Illumina MiSeqDx platform. With the use of the TruSeq amplicon workflow, both strands of DNA from the starting material were interrogated independently. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Nitin Udar, Anita Iyer, Margaret Porter, Robert Haigis, Shannon Smith, Shivani Dhillon, Kristen Meier, Diane Ward, Jing Lu, Paul Wenz, Leonard Buchner, Tamsen Dunn, Aaron Wise, Amy Mueller, Karen Gutekunst Tags: Regular article Source Type: research

Multi-Institutional Evaluation of Interrater Agreement of Variant Classification Based on the 2017 Association for Molecular Pathology, American Society of Clinical Oncology, and College of American Pathologists Standards and Guidelines for the Interpretation and Reporting of Sequence Variants in Cancer
This multi-institutional study was undertaken to evaluate interrater reliability of the 2017 Association for Molecular Pathology/American Society of Clinical Oncology/College of American Pathologists guidelines for interpretation and reporting of oncology sequence variants and to assess current practices and perceptions surrounding these guidelines. Fifty-one variants were distributed to 20 participants from 10 institutions for classification using the new guidelines. Agreement was assessed using chance-corrected agreement (Cohen κ). (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Deepika Sirohi, Robert L. Schmidt, Dara L. Aisner, Amir Behdad, Bryan L. Betz, Noah Brown, Joshua F. Coleman, Christopher L. Corless, Georgios Deftereos, Mark D. Ewalt, Helen Fernandes, Susan J. Hsiao, Mahesh M. Mansukhani, Sarah S. Murray, Nifang Niu, La Tags: Regular article Source Type: research

Digital Rolling Circle Amplification –Based Detection of Ebola and Other Tropical Viruses
Emerging tropical viruses have caused serious outbreaks during the recent years, such as Ebola virus (EBOV) in 2014 and the most recent in 2018 to 2019 in Congo. Thus, immediate diagnostic attention is demanded at the point of care in resource-limited settings, because the performance and the operational parameters of conventional EBOV testing are limited. Especially, their sensitivity, specificity, and coverage of other tropical disease viruses make them unsuitable for diagnostic at the point of care. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Sibel Ciftci, Felix Neumann, Samir Abdurahman, K. Sofia Appelberg, Ali Mirazimi, Mats Nilsson, Narayanan Madaboosi Tags: Regular article Source Type: research

Retrospective validation of a metagenomic sequencing protocol for combined detection of rna and dna viruses using respiratory samples from pediatric patients
In this study, the performance of an in-house mNGS protocol for routine diagnostics of viral respiratory infections with potential for automated pan-pathogen detection was studied.The sequencing protocol and bioinformatics analysis were designed and optimized including exogenous internal controls. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Sander van Boheemen, Anneloes L. van Rijn, Nikos Pappas, Ellen C. Carbo, Ruben H.P. Vorderman, Igor Sidorov, Peter J. van `t Hof, Hailiang Mei, Eric C.J. Claas, Aloys C.M. Kroes, Jutte J.C. de Vries Source Type: research

Therapeutic Monitoring of Circulating DNA Mutations in Metastatic Cancer with Personalized Digital PCR
The objective of this study was to validate the use of personalized dPCR mutation assays to monitor patients with metastatic cancer. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Christina M. Wood-Bouwens, Derrick Haslem, Bryce Moulton, Alison F. Almeda, Hojoon Lee, Gregory M. Heestand, Lincoln D. Nadauld, Hanlee P. Ji Source Type: research

Development and validation of a novel and rapid molecular detection method for high-risk HPV in formalin-fixed paraffin-embedded tumor tissue
In this study, we developed and validated a molecular HPV detection method for FFPE specimens of oropharyngeal and non-oropharyngeal HNSCC. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Steven W. Mes, Dani ëlle A.M. Heideman, Elisabeth Bloemena, Arjen Brink, Martijn Bogaarts, C. René Leemans, Ruud H. Brakenhoff Source Type: research

Multi-Institutional Evaluation of Inter-rater Agreement of Variant Classification Based on the 2017 AMP, ASCO and CAP Standards and Guidelines for the Interpretation and Reporting of Sequence Variants in Cancer
This multi-institutional study was undertaken to evaluate inter-rater reliability of the 2017 AMP/ASCO/CAP guidelines for interpretation and reporting of oncology sequence variants and to assess current practices and perceptions surrounding these guidelines. Fifty-one variants were distributed to 20 participants from 10 institutions for classification using the new guidelines. Agreement was assessed using chance corrected agreement (Cohen ’s kappa). Overall chance corrected agreement (kappa) was 0.35. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Deepika Sirohi, Robert L. Schmidt, Dara L. Aisner, Amir Behdad, Bryan L. Betz, Noah Brown, Joshua F. Coleman, Christopher L. Corless, Georgios Deftereos, Mark D. Ewalt, Helen Fernandes, Susan J. Hsiao, Mahesh M. Mansukhani, Sarah S. Murray, Nifang Niu, La Source Type: research

Sample Tracking Using Unique Sequence Controls
Sample tracking and identity is essential when processing multiple samples in parallel. Sequencing applications often involve high sample numbers and the data is frequently used in a clinical setting. As such a simple and accurate intrinsic sample tracking process through a sequencing pipeline is essential. Various solutions have been implemented to verify sample identity including variant detection at the start and end of the pipeline using arrays or genotyping, bioinformatic comparisons, and optical barcoding of samples. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Richard A. Moore, Thomas Zeng, T. Roderick Docking, Ian Bosdet, Yaron S. Butterfield, Sarah Munro, Irene Li, Lucas Swanson, Elizabeth R. Starks, Kane Tse, Andrew J. Mungall, Robert A. Holt, Aly Karsan Tags: Technical Advance Source Type: research

Level of seven neuroblastoma-associated mRNAs detected by droplet digital PCR is associated with tumor relapse/regrowth of high-risk neuroblastoma patients
Monitoring of several sets of neuroblastoma-associated mRNAs (NB-mRNAs) by quantitative PCR (qPCR) can be used for evaluating minimal residual disease (MRD) in neuroblastoma (NB) patients. Droplet digital PCR (ddPCR) is an adaption of qPCR that potentially provides more simple and reproducible detection of low-level of mRNAs. However, it remains tested whether MRD in NB patients can be monitored by ddPCR using a set of NB-mRNAs. In the present study, 208 bone marrow (BM) and 67 peripheral blood (PB) samples were retrospectively collected from 20 high-risk NB patients with clinical disease evaluation at two Japanese centers...
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Khin Kyae Mon Thwin, Toshiaki Ishida, Suguru Uemura, Nobuyuki Yamamoto, Kyaw San Lin, Akihiro Tamura, Aiko Kozaki, Atsuro Saito, Kenji Kishimoto, Takeshi Mori, Daiichiro Hasegawa, Yoshiyuki Kosaka, Nanako Nino, Satoru Takafuji, Kazumoto Iijima, Noriyuki N Tags: Regular Article Source Type: research

Concordance of Genomic Alterations by Next-generation Sequencing (NGS) in Tumor Tissue vs. Cell-free DNA in Stage I –IV Non-small Cell Lung Cancer
Molecular biomarkers hold promise for the personalization of cancer treatment. However, a typical tumor biopsy may be difficult to acquire and may not capture genetic variations within or across tumors. Given these limitations, tumor genotyping using next-generation sequencing (NGS) of plasma-derived circulating tumor DNA (ctDNA) has the potential to transform non-small cell lung cancer (NSCLC) management. Importantly, mutations detected in biopsied tissue must also be detected in plasma-derived ctDNA at different disease stages. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: John Jiang, Hans-Peter Adams, Lijing Yao, Stephanie Yaung, Preeti Lal, Aarthi Balasubramanyam, Frederike Fuhlbr ück, Nalin Tikoo, Alexander F. Lovejoy, Sebastian Froehler, Li Tai Fang, H. Jost Achenbach, Ralph Floegel, Rainer Krügel, John Palma Tags: Regular Article Source Type: research

Development and Analytical Validation of a DNA Dual-Strand Approach for the FDA-Approved Next-Generation Sequencing-Based Praxis Extended RAS Panel for Metastatic Colorectal Cancer Samples
A next-generation sequencing method was developed that can distinguish single-stranded modifications from low-frequency somatic mutations present on both strands of DNA in formalin-fixed paraffin-embedded (FFPE) colorectal cancer (CRC) samples. We applied this method for analytical validation of the Praxis Extended RAS Panel, a United States Food and Drug Administration (FDA)-approved companion diagnostic for panitumumab, on the Illumina MiSeqDx platform. Using the TruSeq amplicon workflow, both strands of DNA from the starting material were interrogated independently. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Nitin Udar, Anita Iyer, Margaret Porter, Robert Haigis, Shannon Smith, Shivani Dhillon, Kristen Meier, Diane Ward, Jing Lu, Paul Wenz, Leonard Buchner, Tamsen Dunn, Aaron Wise, Amy Mueller, Karen Gutekunst Source Type: research

Digital rolling circle amplification-based detection of Ebola and other tropical viruses
Emerging tropical viruses have caused serious outbreaks during the recent years, such as Ebola virus (EBOV) in 2014 and the most recent in 2018-19 in Congo. Thus, immediate diagnostic attention is demanded at the point-of-care in resource-limited settings, since the performance and the operational parameters of conventional EBOV testing are limited. Especially, their sensitivity, specificity and coverage of other tropical disease viruses make them unsuitable for diagnostic at the point-of-care. Here, we present a padlock probe (PLP)-based rolling circle amplification (RCA) method for the detection of EBOV. (Source: Journal...
Source: Journal of Molecular Diagnostics - December 11, 2019 Category: Pathology Authors: Sibel Ciftci, Felix Neumann, Samir Abdurahman, K. Sofia Appelberg, Ali Mirazimi, Mats Nilsson, Narayanan Madaboosi Source Type: research

Clinical Validation of a Myeloid Next-Generation Sequencing Panel for Single-Nucleotide Variants, Insertions/Deletions, and Fusion Genes
Myeloid neoplasms are a heterogeneous group of neoplasms including acute myeloid leukemia (AML), myeloproliferative neoplasms, myelodysplastic syndrome, and myeloproliferative neoplasms/myelodysplastic syndrome. Genetic abnormalities are used as diagnostic, prognostic, and predictive biomarkers in patients with these diseases. Herein, we describe the clinical validation of the Oncomine Myeloid Research (OMR) next-generation sequencing panel that interrogates for 40 genes and 29 fusion genes commonly seen in myeloid neoplasms. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Iyare Izevbaye, Li Y. Liang, Cheryl Mather, Soufiane El-Hallani, Remegio Maglantay, Lalit Saini Tags: Regular article Source Type: research

VarCover
To facilitate reference-material selection for clinical genetic testing laboratories, we developed VarCover, open-source software hosted on GitHub, which accepts a file of variants (rsIDs or VCF) and returns an approximately minimal set of samples covering the targeted alleles. VarCover employs the SetCoverPy package, sample weights, and preselection of singleton-possessing samples to efficiently solve the min-set cover problem. As a test case, we attempted to find a minimal set of reference samples from the 1000 Genomes Project to cover 237 variants considered putatively pathogenic (of which 12 were classified as pathogen...
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Erick R. Scott, Vikas Bansal, Carl Meacham, Stuart A. Scott Tags: Technical advance Source Type: research

Subgroup Analysis Can Optimize the Relapse-Prediction Cutoff Value for WT1 Expression After Allogeneic Hematologic Stem Cell Transplantation in Acute Myeloid Leukemia
High WT1 expression after allogeneic hematologic stem cell transplantation (allo-HSCT) can strongly predict relapse in acute myeloid leukemia (AML). However, the cutoff values obtained have been inconsistent. Precise cutoff values may be optimized through subtype analysis; the RUNX1-RUNX1T1 fusion transcript provides an ideal reference. RUNX1-RUNX1T1 and WT1 transcript levels were simultaneously measured in 1299 bone marrow samples serially collected from 176 t(8;21) AML patients after allo-HSCT. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Ya-Zhen Qin, Yu Wang, Lan-Ping Xu, Xiao-Hui Zhang, Xiao-Su Zhao, Kai-Yan Liu, Xiao-Jun Huang Tags: Regular article Source Type: research

Digital Gene Expression Analysis on Cytology  Smears Can Rule Out Malignancy in Follicular-Patterned Thyroid Tumors
Patients with indeterminate thyroid nodules (Bethesda III and IV) are often treated with diagnostic lobectomy, which in most cases represents an overtreatment. A reliable rule-out molecular test could spare patients unnecessary surgery. Stained smears of 88 indeterminate thyroid nodules with histologic diagnosis of follicular-patterned tumors were selected: 34 follicular adenomas (FAs), 34 follicular variant papillary thyroid carcinomas (FVPTCs), and 20 noninvasive follicular neoplasms with papillary-like nuclear features (NIFTPs). (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Elisabetta Macerola, Anello Marcello Poma, Agnese Proietti, Rossana Romani, Liborio Torregrossa, Clara Ugolini, Teresa Rago, Paolo Vitti, Fulvio Basolo Source Type: research

Single-Molecule Sequencing
We investigated the potential of next-generation sequencing (NGS) as an alternative method for preimplantation genetic testing of monogenic disease (PGT-M) with human leukocyte antigen (HLA) matching and for noninvasive prenatal diagnosis follow-up. The case involved parents who were carriers of the FANCG 260delG mutation. After clinical PGT using conventional STR and mutation analysis, two euploid disease-free embryos were transferred, resulting in a twin pregnancy. Using the original embryo whole genome amplification products from 10 embryos, NGS confirmed the genotypes of the eight nontransferred embryos for both mutati...
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Svetlana Rechitsky, Anver Kuliev, Geraldine San Ramon, Ilan Tur-Kaspa, Yin Wang, Wenjie Wang, Xueqing Wu, Li Wang, Don Leigh, David S. Cram Tags: Regular article Source Type: research

Clinical Validation of a Myeloid Next-Generations Sequencing Panel for Single-Nucleotide Variants, Insertions/Deletions, and Fusion Genes
Myeloid neoplasms are a heterogeneous group of neoplasms including acute myeloid leukemia (AML), myeloproliferative neoplasms, myelodysplastic syndrome, and myeloproliferative neoplasms/myelodysplastic syndrome. Genetic abnormalities are used as diagnostic, prognostic, and predictive biomarkers in patients with these diseases. Herein, we describe the clinical validation of the Oncomine Myeloid Research (OMR) next-generation sequencing panel that interrogates for 40 genes and 29 fusion genes commonly seen in myeloid neoplasms. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Iyare Izevbaye, Li Ying Liang, Cheryl Mather, Soufiane El-Hallani, Remegio Maglantay, Lalit Saini Tags: Regular article Source Type: research

Single Molecule Sequencing A New Approach for Preimplantation Testing and Noninvasive Prenatal Diagnosis Confirmation of Fetal Genotype
We investigated the potential of next-generation sequencing (NGS) as an alternative methodology for preimplantation genetic testing of monogenic disease (PGT-M) with HLA matching and for noninvasive prenatal diagnosis follow up. The case involved parents who were carriers of the FANCG 260delG mutation. Following clinical PGT using conventional STR and mutation analysis, two euploid disease-free embryos were transferred resulting in a twin pregnancy. Using the original embryo whole genome amplification products from 10 embryos, NGS confirmed the genotypes of the eight non-transferred embryos for both mutation status and HLA...
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Svetlana Rechitsky, Anver Kuliev, Geraldine San Ramon, Ilan Tur-Kaspa, Yin Wang, Wenjie Wang, Xueqing Wu, Li Wang, Don Leigh, David S. Cram Tags: Regular article Source Type: research

Clinical Validation of a Myeloid Next-Generations Sequencing Panel for Single Nucleotide Variants, Indels, and Fusion Genes
Myeloid neoplasms are a heterogenous group of neoplasms including acute myeloid leukemia (AML), myeloproliferative neoplasms, myelodysplastic syndrome, and myeloproliferative neoplasms / myelodysplastic syndrome. Genetic abnormalities are used as diagnostic, prognostic, and predictive biomarkers in patients with these diseases. Here, we describe the clinical validation of the Oncomine Myeloid Research (OMR) next-generation sequencing panel that interrogates for 40 genes and 29 fusion genes commonly seen in myeloid neoplasms. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Iyare Izevbaye, Li Ying Liang, Cheryl Mather, Soufiane El-Hallani, Remegio Maglantay JR, Lalit Saini Tags: Regular Article Source Type: research

Subgroup Analysis Can Optimize the Relapse Prediction Cutoff Value for WT1 Expression Following Allogeneic Hematologic Stem Cell Transplantation in Acute Myeloid Leukemia
High WT1 expression after allogeneic hematologic stem cell transplantation (allo-HSCT) can strongly predict relapse in acute myeloid leukemia (AML). However, the cutoff values obtained were inconsistent. The precise cutoff values may be optimized through subtype analysis, and the RUNX1-RUNX1T1 fusion transcript provides an ideal reference. RUNX1-RUNX1T1 and WT1 transcript levels were simultaneously measured in 1,299 bone marrow samples serially collected from 176 t(8;21) AML patients after receiving allo-HSCTfor. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Ya-Zhen Qin, Yu Wang, Lan-Ping Xu, Xiao-Hui Zhang, Xiao-Su Zhao, Kai-Yan Liu, Xiao-Jun Huang Tags: Regular Article Source Type: research

Evaluation of the BD MAX Check-Points CPO assay for the detection of carbapenemase producers directly from rectal swabs
A novel real time multiplex PCR assay, BD MAX Check-Points CPO, was evaluated to detect carbapenemase-producing organisms in clinical settings on the BD MAX system. A total of 175 well-characterized isolates (including 123 carbapenemase-producers) and 128 rectal swab specimens (including 83 positives) of patients considered at “high-risk” for carriage of carbapenemase-producers were included. Bacterial suspensions were used to spike “true negative” rectal swabs to mimic a clinical sample. Fifty microliters of sample, containing either the spiked or the patient’s samples were processed. (Source...
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Delphine Girlich, Saoussen Oueslati, Sandrine Bernabeu, Isabelle Langlois, Christine Begasse, Nicolas Arangia, Elodie Creton, Garance Cotellon, Aimie Sauvadet, Laurent Dortet, Nicolas Fortineau, Thierry Naas Tags: Regular Article Source Type: research

Evaluation of Commercial Next-Generation Sequencing Bioinformatics Software Solutions
Next-generation sequencing (NGS) diagnostics continue to expand rapidly in clinical medicine. An ever-expanding menu of molecular biomarkers are deemed important for diagnostic, prognostic, and therapeutic assessment in patients. The increasing role of NGS in the clinic is driven mainly by the falling costs of sequencing. However, the data-intensive nature of NGS makes bioinformatic analysis a major challenge to many clinical laboratories. Critically needed NGS bioinformatics personnel are hard to recruit and retain in small- to mid-size clinical laboratories. (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Rama R. Gullapalli Source Type: research

Digital Gene Expression Analysis on Cytology Smears Can Rule-out Malignancy in Follicular-Patterned Thyroid Tumors
Patients with indeterminate thyroid nodules (Bethesda III and IV) are often treated with diagnostic lobectomy, which in the majority of cases represent an overtreatment. A reliable rule-out molecular test could spare patients unnecessary surgery. Stained smears of 88 indeterminate thyroid nodules with histological diagnosis of follicular-patterned tumors were selected: 34 follicular adenomas (FA), 34 follicular variant papillary thyroid carcinomas (FVPTC), and 20 noninvasive follicular neoplasms with papillary-like nuclear features (NIFTP). (Source: Journal of Molecular Diagnostics)
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Elisabetta Macerola, Anello Marcello Poma, Agnese Proietti, Rossana Romani, Liborio Torregrossa, Clara Ugolini, Teresa Rago, Paolo Vitti, Fulvio Basolo Source Type: research

VarCover: Allele Min-Set Cover Software
To facilitate reference material selection for clinical genetic testing laboratories, we developed VarCover, open-source software hosted on GitHub, which accepts a file of variants (rsIDs or VCF) and returns an approximately minimal set of samples covering the targeted alleles. VarCover employs the SetCoverPy package, sample weights, and pre-selection of singleton-possessing samples to efficiently solve the min-set cover problem. As a test case, we attempted to find a minimal set of 1000 Genomes (1KG) Project reference samples to cover 237 putatively pathogenic variants (of which 12 were pathogenic or likely pathogenic) in...
Source: Journal of Molecular Diagnostics - November 18, 2019 Category: Pathology Authors: Erick R. Scott, Vikas Bansal, Carl Meacham, Stuart A. Scott Tags: Technical Advance Source Type: research