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A rapid, multiplex digital PCR assay to detect gene variants and fusions in non‐small cell lung cancer

Digital PCR (dPCR) is emerging as an ideal platform for the detection and tracking of genomic variants in cancer due to its high sensitivity and simple workflow. The growing number of clinically actionable cancer biomarkers creates a need for fast, accessible methods that allow for dense information...

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Autores principales: Leatham, Bryan, McNall, Katie, Subramanian, Hari K. K., Jacky, Lucien, Alvarado, John, Yurk, Dominic, Wang, Mimi, Green, Donald C., Tsongalis, Gregory J., Rajagopal, Aditya, Schwartz, Jerrod J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10620117/
https://www.ncbi.nlm.nih.gov/pubmed/37714814
http://dx.doi.org/10.1002/1878-0261.13523
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author Leatham, Bryan
McNall, Katie
Subramanian, Hari K. K.
Jacky, Lucien
Alvarado, John
Yurk, Dominic
Wang, Mimi
Green, Donald C.
Tsongalis, Gregory J.
Rajagopal, Aditya
Schwartz, Jerrod J.
author_facet Leatham, Bryan
McNall, Katie
Subramanian, Hari K. K.
Jacky, Lucien
Alvarado, John
Yurk, Dominic
Wang, Mimi
Green, Donald C.
Tsongalis, Gregory J.
Rajagopal, Aditya
Schwartz, Jerrod J.
author_sort Leatham, Bryan
collection PubMed
description Digital PCR (dPCR) is emerging as an ideal platform for the detection and tracking of genomic variants in cancer due to its high sensitivity and simple workflow. The growing number of clinically actionable cancer biomarkers creates a need for fast, accessible methods that allow for dense information content and high accuracy. Here, we describe a proof‐of‐concept amplitude modulation‐based multiplex dPCR assay capable of detecting 12 single‐nucleotide and insertion/deletion (indel) variants in EGFR, KRAS, BRAF, and ERBB2, 14 gene fusions in ALK, RET, ROS1, and NTRK1, and MET exon 14 skipping present in non‐small cell lung cancer (NSCLC). We also demonstrate the use of multi‐spectral target‐signal encoding to improve the specificity of variant detection by reducing background noise by up to an order of magnitude. The assay reported an overall 100% positive percent agreement (PPA) and 98.5% negative percent agreement (NPA) compared with a sequencing‐based assay in a cohort of 62 human formalin‐fixed paraffin‐embedded (FFPE) samples. In addition, the dPCR assay rescued actionable information in 10 samples that failed to sequence, highlighting the utility of a multiplexed dPCR assay as a potential reflex solution for challenging NSCLC samples.
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spelling pubmed-106201172023-11-03 A rapid, multiplex digital PCR assay to detect gene variants and fusions in non‐small cell lung cancer Leatham, Bryan McNall, Katie Subramanian, Hari K. K. Jacky, Lucien Alvarado, John Yurk, Dominic Wang, Mimi Green, Donald C. Tsongalis, Gregory J. Rajagopal, Aditya Schwartz, Jerrod J. Mol Oncol Method Digital PCR (dPCR) is emerging as an ideal platform for the detection and tracking of genomic variants in cancer due to its high sensitivity and simple workflow. The growing number of clinically actionable cancer biomarkers creates a need for fast, accessible methods that allow for dense information content and high accuracy. Here, we describe a proof‐of‐concept amplitude modulation‐based multiplex dPCR assay capable of detecting 12 single‐nucleotide and insertion/deletion (indel) variants in EGFR, KRAS, BRAF, and ERBB2, 14 gene fusions in ALK, RET, ROS1, and NTRK1, and MET exon 14 skipping present in non‐small cell lung cancer (NSCLC). We also demonstrate the use of multi‐spectral target‐signal encoding to improve the specificity of variant detection by reducing background noise by up to an order of magnitude. The assay reported an overall 100% positive percent agreement (PPA) and 98.5% negative percent agreement (NPA) compared with a sequencing‐based assay in a cohort of 62 human formalin‐fixed paraffin‐embedded (FFPE) samples. In addition, the dPCR assay rescued actionable information in 10 samples that failed to sequence, highlighting the utility of a multiplexed dPCR assay as a potential reflex solution for challenging NSCLC samples. John Wiley and Sons Inc. 2023-10-04 /pmc/articles/PMC10620117/ /pubmed/37714814 http://dx.doi.org/10.1002/1878-0261.13523 Text en © 2023 ChromaCode and The Authors. Molecular Oncology published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies. https://creativecommons.org/licenses/by/4.0/This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Method
Leatham, Bryan
McNall, Katie
Subramanian, Hari K. K.
Jacky, Lucien
Alvarado, John
Yurk, Dominic
Wang, Mimi
Green, Donald C.
Tsongalis, Gregory J.
Rajagopal, Aditya
Schwartz, Jerrod J.
A rapid, multiplex digital PCR assay to detect gene variants and fusions in non‐small cell lung cancer
title A rapid, multiplex digital PCR assay to detect gene variants and fusions in non‐small cell lung cancer
title_full A rapid, multiplex digital PCR assay to detect gene variants and fusions in non‐small cell lung cancer
title_fullStr A rapid, multiplex digital PCR assay to detect gene variants and fusions in non‐small cell lung cancer
title_full_unstemmed A rapid, multiplex digital PCR assay to detect gene variants and fusions in non‐small cell lung cancer
title_short A rapid, multiplex digital PCR assay to detect gene variants and fusions in non‐small cell lung cancer
title_sort rapid, multiplex digital pcr assay to detect gene variants and fusions in non‐small cell lung cancer
topic Method
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10620117/
https://www.ncbi.nlm.nih.gov/pubmed/37714814
http://dx.doi.org/10.1002/1878-0261.13523
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