Aims: Gene fusions assays are key for personalised treatments of advanced human cancers. Their implementation on cytological material requires a preliminary validation that may make use of cell line slides mimicking cytological samples. In this international multi-institutional study, gene fusion reference standards were developed and validated. Methods: Cell lines harbouring EML4(13)-ALK(20) and SLC34A2(4)-ROS1(32) gene fusions were adopted to prepare reference standards. Eight laboratories (five adopting amplicon-based and three hybridisation-based platforms) received, at different dilution points two sets of slides (slide A 50.0%, slide B 25.0%, slide C 12.5% and slide D wild type) stained by Papanicolaou (Pap) and May Grunwald Giemsa (MGG). Analysis was carried out on a total of 64 slides. Results: Four (50.0%) out of eight laboratories reported results on all slides and dilution points. While 12 (37.5%) out of 32 MGG slides were inadequate, 27 (84.4%) out of 32 Pap slides produced libraries adequate for variant calling. The laboratories using hybridisation-based platforms showed the highest rate of inadequate results (13/24 slides, 54.2%). Conversely, only 10.0% (4/40 slides) of inadequate results were reported by laboratories adopting amplicon-based platforms. Conclusions: Reference standards in cytological format yield better results when Pap staining and processed by amplicon-based assays. Further investigation is required to optimise these standards for MGG stained cells and for hybridisation-based approaches.

Reference standards for gene fusion molecular assays on cytological samples: an international validation study

de Biase, Dario;Tallini, Giovanni;
2023

Abstract

Aims: Gene fusions assays are key for personalised treatments of advanced human cancers. Their implementation on cytological material requires a preliminary validation that may make use of cell line slides mimicking cytological samples. In this international multi-institutional study, gene fusion reference standards were developed and validated. Methods: Cell lines harbouring EML4(13)-ALK(20) and SLC34A2(4)-ROS1(32) gene fusions were adopted to prepare reference standards. Eight laboratories (five adopting amplicon-based and three hybridisation-based platforms) received, at different dilution points two sets of slides (slide A 50.0%, slide B 25.0%, slide C 12.5% and slide D wild type) stained by Papanicolaou (Pap) and May Grunwald Giemsa (MGG). Analysis was carried out on a total of 64 slides. Results: Four (50.0%) out of eight laboratories reported results on all slides and dilution points. While 12 (37.5%) out of 32 MGG slides were inadequate, 27 (84.4%) out of 32 Pap slides produced libraries adequate for variant calling. The laboratories using hybridisation-based platforms showed the highest rate of inadequate results (13/24 slides, 54.2%). Conversely, only 10.0% (4/40 slides) of inadequate results were reported by laboratories adopting amplicon-based platforms. Conclusions: Reference standards in cytological format yield better results when Pap staining and processed by amplicon-based assays. Further investigation is required to optimise these standards for MGG stained cells and for hybridisation-based approaches.
2023
Malapelle, Umberto; Pepe, Francesco; Pisapia, Pasquale; Altimari, Annalisa; Bellevicine, Claudio; Brunnström, Hans; Bruno, Rossella; Büttner, Reinhard; Cirnes, Luis; De Andrea, Carlos E; de Biase, Dario; Dumur, Catherine I; Ericson Lindquist, Kajsa; Fontanini, Gabriella; Gautiero, Eugenio; Gentien, David; Hofman, Paul; Hofman, Veronique; Iaccarino, Antonino; Lozano, Maria Dolores; Mayo-de-Las-Casas, Clara; Merkelbach-Bruse, Sabine; Pagni, Fabio; Roman, Ruth; Schmitt, Fernando C; Siemanowski, Janna; Roy-Chowdhuri, Sinchita; Tallini, Giovanni; Tresserra, Francesc; Vander Borght, Sara; Vielh, Philippe; Vigliar, Elena; Vita, Giulia Anna Carmen; Weynand, Birgit; Rosell, Rafael; Molina Vila, Miguel Angel; Troncone, Giancarlo
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11585/839994
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