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TargetPlex FFPE-Direct DNA Library Preparation Kit for SiRe NGS panel: an international performance evaluation study
  1. Umberto Malapelle1,
  2. Francesco Pepe1,
  3. Pasquale Pisapia1,
  4. Roberta Sgariglia1,
  5. Mariantonia Nacchio1,
  6. Massimo Barberis2,
  7. Michel Bilh3,
  8. Lukas Bubendorf3,
  9. Reinhard Büttner4,
  10. Daniela Cabibi5,
  11. Marta Castiglia6,
  12. Carlos E De Andrea7,
  13. Dario de Biase8,
  14. Catherine I Dumur9,
  15. Gabriella Fontanini10,
  16. Javier Freire11,
  17. Valerio Gristina6,
  18. Paul Hofman12,
  19. Marius Ilie13,
  20. Maria Dolores Lozano14,
  21. Sabine Merkelbach-Bruse4,
  22. Roberto Pappesch4,
  23. Natalie Pelusi15,
  24. Gianluca Roma16,
  25. Antonio Russo6,
  26. Spasenija Savic3,
  27. Janna Siemanowski4,
  28. Giovanni Tallini17,
  29. Verena Tischler15,
  30. Sara Vander Borght18,
  31. Birgit Weynand18,
  32. Tom Xu19,
  33. Giancarlo Troncone1
  1. 1 Public Health, University of Naples Federico II, Naples, Italy
  2. 2 Clinic Unit of Histopathology and Molecular Diagnostics, Istituto Europeo di Oncologia, Milano, Italy
  3. 3 Department of Pathology, University Hospital Basel, Basel, Switzerland
  4. 4 Department of Pathology, University of Cologne, Cologne, Germany
  5. 5 Health Promotion Sciences, Maternal and Infantile Care, Internal Medicine and Medical Specialties, University of Palermo, Palermo, Italy
  6. 6 Surgical, Oncological and Oral Sciences, University of Palermo, Palermo, Italy
  7. 7 Pathology, University of Navarra, Pamplona, Spain
  8. 8 Medicine (DIMES)a Hospital, Anatomic Pathology Unit, University of Bologna, Bologna, Italy
  9. 9 Molecular Diagnostic Department, Aurora Diagnostics, Jacksonville, Florida, USA
  10. 10 Surgical, Medical, Molecular Pathology and Critical Area, University of Pisa, Pisa, Italy
  11. 11 Pathology, Hospital Universitario Marques de Valdecilla, Santander, Spain
  12. 12 Pathology, INSERM, Nice, France
  13. 13 Laboratory of Clinical and Experimental Pathology, Université Côte d'Azur, Nice, France
  14. 14 Pathology, Universidad de Navarra-Clínica Universidad de Navarra, Pamplona, Spain
  15. 15 Pathology, University of Bonn, Bonn, Germany
  16. 16 R&D Department, TargetPlex Genomics, Belmont, California, USA
  17. 17 Pathology, University of Bologna, Bologna, Italy
  18. 18 Department of Pathology, Katholieke Universiteit Leuven UZ Leuven, Leuven, Belgium
  19. 19 R&D Department, SenseCare Medicals, Inc, Pleasanton, California, USA
  1. Correspondence to Professor Giancarlo Troncone, Public Health, University of Naples Federico II, Naples, Italy; giancarlo.troncone{at}


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
  • pathology
  • molecular
  • lung neoplasms
  • biomarkers
  • tumour

Data availability statement

All data relevant to the study are included in the article or uploaded as supplemental information.

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Data availability statement

All data relevant to the study are included in the article or uploaded as supplemental information.

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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.

  • Provenance and peer review Not commissioned; internally peer reviewed.

  • Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.