Aim Next generation sequencing (NGS) represents a key diagnostic tool to identify clinically relevant gene alterations for treatment-decision making in cancer care. However, the complex manual workflow required for NGS has limited its implementation in routine clinical practice. In this worldwide study, we validated the clinical performance of the TargetPlex FFPE-Direct DNA Library Preparation Kit for NGS analysis. Impressively, this new assay obviates the need for separate, labour intensive and time-consuming pre-analytical steps of DNA extraction, purification and isolation from formalin-fixed paraffin embedded (FFPE) specimens in the NGS workflow.
Methods The TargetPlex FFPE-Direct DNA Library Preparation Kit, which enables NGS analysis directly from FFPE, was specifically developed for this study by TargetPlex Genomics Pleasanton, California. Eleven institutions agreed to take part in the study coordinated by the Molecular Cytopathology Meeting Group (University of Naples Federico II, Naples, Italy). All participating institutions received a specific Library Preparation Kit to test eight FFPE samples previously assessed with standard protocols. The analytical parameters and mutations detected in each sample were then compared with those previously obtained with standard protocols.
Results Overall, 92.8% of the samples were successfully analysed with the TargetPlex FFPE-Direct DNA Library Preparation Kit on Thermo Fisher Scientific and Illumina platforms. Altogether, in comparison with the standard workflow, the TargetPlex FFPE-Direct DNA Library Preparation Kit was able to detect 90.5% of the variants.
Conclusion The TargetPlex FFPE-Direct DNA Library Preparation Kit combined with the SiRe panel constitutes a convenient, practical and robust cost-saving solution for FFPE NGS analysis in routine practice.
- molecular biology
- lung neoplasms
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Handling editor Runjan Chetty.
Twitter @UmbertoMalapel1, @PasqualePisapia
Contributors Conceptualisation: UM and GTr. Methodology: all authors. Software: all authors. Validation: all authors. Formal analysis: all authors. Investigation: all authors. Resources: all authors. Data curation: all authors. Writing—original draft preparation: FP, PP, UM, GTr. Writing—review and editing: all authors. Visualisation: all authors. Supervision: UM and GTr. Project administration: UM and GTr. Funding acquisition: GTr.
Funding Monitoraggio ambientale, studio ed approfondimento della salute della popolazione residente in aree a rischio—In attuazione della D.G.R. Campanian.180/2019. POR Campania FESR 2014–2020 Progetto ‘Sviluppo di Approcci Terapeutici Innovativi per patologie Neoplastiche resistenti ai trattamenti—SATIN’.
Competing interests UM has received personal fees (as consultant and/or speaker bureau) from Boehringer Ingelheim, Roche, MSD, Amgen, Thermo Fisher Scientifics, Diaceutics, GSK, Merck and AstraZeneca, unrelated to the current work. LB is has a consulting or advisory role with AstraZeneca, AbbVie, Bayer, Boehringer Ingelheim, Eli Lilly, MSD, Pfizer, Takeda and F. Hoffmann-La Roche, and has received research funding (institution) from F. Hoffmann-La Roche, MSD and Sanofi. PH reports personal fees (as advisor) from Roche, Astrazeneca, BMS, MSD, Pfizer, Bayer, Amgen, Illumina, Qiagen, Thermo Fisher Scientific, Biocartis, Ed Lilly, unrelated to the current work. MI reports personal fees (as speaker bureau) from Roche, Merck & Co, AstraZeneca, Bristol-Myers Squibb and Boehringer-Ingelheim, unrelated to the current work. GR is the coinventor of the intellectual property used in the background noise suppression aspects of the TargePlex FFPE-Direct NGS Library Preparation Kit developed in collaboration with SenseCare Medicals. AR reports personal fees from Bristol, Pfizer, Bayer, Kyowa Kirin, Ambrosetti for advisory board activity and speaker honorarium from Roche Diagnostic speaker honorarium, unrelated to the submitted work. SS received personal fees from MSD, Astra Zeneca, Boehringer Ingelheim, Roche, Pfizer, Bristol-Myers Squibb and Thermo Fisher Scientific, unrelated to the submitted work. TX is an employee and affiliate with SenseCare Medicals. GTroncone reports personal fees (as speaker bureau or advisor) from Roche, MSD, Pfizer and Bayer, unrelated to the current work.The other authors have nothing to disclose.
Patient consent for publication Not required.
Ethics approval All the experimental procedures were carried out in accordance with the general authorisation to process personal data for scientific research purposes from ‘The Italian Data Protection Authority’ (http://www.garanteprivacy.it/web/guest/home/docweb/-/docwebdisplay/export/2485392). All information regarding human material was managed using anonymous numerical codes, and all samples were handled in compliance with the Declaration of Helsinki (https://www.wma.net/fr/news-post/en-matierede-transfert-des-taches-la-securite-des-patients-et-la-qualitedes-soins-devraient-etre-primordiales/). According to the aforementioned national guidelines, the double-blind study did not require an Ethical Committee approval since it did not affect the clinical management of patient samples.
Provenance and peer review Not commissioned; internally peer reviewed.
Data availability statement All data relevant to the study are included in the article or uploaded as supplemental information.
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