96 results on '"Gragnano, Gianluca"'
Search Results
2. Next generation sequencing for liquid biopsy based testing in non-small cell lung cancer in 2021
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Pisapia, Pasquale, Costa, José Luis, Pepe, Francesco, Russo, Gianluca, Gragnano, Gianluca, Russo, Alessandro, Iaccarino, Antonino, de Miguel-Perez, Diego, Serrano, Maria Josè, Denninghoff, Valeria, Quagliata, Luca, Rolfo, Christian, and Malapelle, Umberto
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- 2021
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3. NGAL Mediates Anaplastic Thyroid Carcinoma Cells Survival Through FAS/CD95 Inhibition
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Crescenzi, Elvira, primary, Mellone, Stefano, additional, Gragnano, Gianluca, additional, Iaccarino, Antonino, additional, Leonardi, Antonio, additional, and Pacifico, Francesco, additional
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- 2023
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4. Liquid biopsy for lung cancer: A cross section on the diagnostic routine experience of a referral Italian institution
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de Luca, Caterina, primary, Russo, Gianluca, additional, Nacchio, Mariantonia, additional, Ingenito, Maria, additional, Palumbo, Lucia, additional, Gragnano, Gianluca, additional, Conticelli, Floriana, additional, Russo, Maria, additional, Rocco, Danilo, additional, Gridelli, Cesare, additional, Bianco, Roberto, additional, Galetta, Domenico, additional, Troncone, Giancarlo, additional, Parente, Paola, additional, and Iaccarino, Antonino, additional
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- 2023
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5. NGAL Mediates Anaplastic Thyroid Carcinoma Cells Survival Through FAS/CD95 Inhibition.
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Crescenzi, Elvira, Mellone, Stefano, Gragnano, Gianluca, Iaccarino, Antonino, Leonardi, Antonio, and Pacifico, Francesco
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LIPOCALIN-2 ,SIDEROPHORES ,THYROID cancer - Abstract
Neutrophil gelatinase-associated lipocalin (NGAL), a siderophore-mediated iron binding protein, is highly expressed in human anaplastic thyroid carcinomas (ATCs) where it plays pleiotropic protumorigenic roles including that of a prosurvival protein. Here we show that NGAL inhibits FAS/CD95 death receptor to control ATC cell survival. FAS/CD95 expression in human specimens from patients with ATC and in ATC-derived cell lines negatively correlate with NGAL expression. Silencing of NGAL in ATC cells leads to FAS/CD95 upregulation, whereas NGAL overexpression determines the opposite effect. As a result, an agonist anti-FAS/CD95 antibody induces cell death in NGAL-silenced cells while it is ineffective on NGAL-overexpressing cells. Interestingly, the inhibitory activity of NGAL on FAS/CD95 is due to its iron carrier property given that perturbing iron homeostasis of NGAL-proficient and -deficient ATC cells directly influences FAS/CD95 expression. Accordingly, conditioned media containing a mutant form of NGAL unable to bind siderophores cannot rescue cells from FAS/CD95-dependent death, whereas NGAL wild type–containing conditioned media abolish the effects of the agonist antibody. We also find that downregulation of FAS/CD95 expression is mediated by iron-dependent NGAL suppression of p53 transcriptional activity. Our results indicate that NGAL contributes to ATC cell survival by iron-mediated inhibition of p53-dependent FAS/CD95 expression and suggest that restoring FAS/CD95 by NGAL suppression could be a helpful strategy to kill ATC cells. [ABSTRACT FROM AUTHOR]
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- 2024
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6. Technical validation of a fully integrated NGS platform in real-world practice of Italian referral institution
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De Luca, Caterina, primary, Pepe, Francesco, additional, Russo, Gianluca, additional, Nacchio, Mariantonia, additional, Pisapia, Pasquale, additional, Russo, Maria, additional, Conticelli, Floriana, additional, Palumbo, Lucia, additional, Scimone, Claudia, additional, Cozzolino, Domencio, additional, Gragnano, Gianluca, additional, Iaccarino, Antonino, additional, Troncone, Giancarlo, additional, and Malapelle, Umberto, additional
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- 2023
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7. Technical Validation of a Fully Integrated NGS Platform in the Real-World Practice of Italian Referral Institutions.
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De Luca, Caterina, Pepe, Francesco, Russo, Gianluca, Nacchio, Mariantonia, Pisapia, Pasquale, Russo, Maria, Conticelli, Floriana, Palumbo, Lucia, Scimone, Claudia, Cozzolino, Domenico, Gragnano, Gianluca, Iaccarino, Antonino, Troncone, Giancarlo, and Malapelle, Umberto
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MOLECULAR diagnosis ,DNA sequencing ,BIOMARKERS ,INDIVIDUALIZED medicine ,MOLECULAR pathology - Abstract
Aims: To date, precision medicine has played a pivotal role in the clinical administration of solid-tumor patients. In this scenario, a rapidly increasing number of predictive biomarkers have been approved in diagnostic practice or are currently being investigated in clinical trials. A pitfall in molecular testing is the diagnostic routine sample available to analyze predictive biomarkers; a scant tissue sample often represents the only diagnostical source of nucleic acids with which to conduct molecular analysis. At the sight of these critical issues, next-generation sequencing (NGS) platforms emerged as referral testing strategies for the molecular analysis of predictive biomarkers in routine practice, but the need for highly skilled personnel and extensive working time drastically impacts the widespread diffusion of this technology in diagnostic settings. Here, we technically validate a fully integrated NGS platform on diagnostic routine tissue samples previously tested with an NGS-based diagnostic workflow by a referral institution. Methods: A retrospective series of n = 64 samples (n = 32 DNA, n = 32 RNA samples), previously tested using a customized NGS assay (SiRe™ and SiRe fusion), was retrieved from the internal archive of the University of Naples Federico II. Each sample was tested by adopting an Oncomine Precision Assay (OPA), which is able to detect 2769 molecular actionable alterations [hotspot mutations, copy number variations (CNV) and gene fusions] on fully integrated NGS platforms (Genexus, Thermo Fisher Scientific (Waltham, MA, USA). The concordance rate between these technical approaches was determined. Results: The Genexus system successfully carried out molecular analysis in all instances. A concordance rate of 96.9% (31 out of 32) was observed between the OPA and SiRe™ panels both for DNA- and RNA-based analysis. A negative predictive value of 100% and a positive predictive value of 96.9% (62 out of 64) were assessed. Conclusions: A fully automatized Genexus system combined with OPA (Thermo Fisher Scientific) may be considered a technically valuable, time-saving sequencing platform to test predictive biomarkers in diagnostic routine practice. [ABSTRACT FROM AUTHOR]
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- 2023
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8. An Italian Multicenter Perspective Harmonization Trial for the Assessment of MET Exon 14 Skipping Mutations in Standard Reference Samples
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Bironzo, Paolo, primary, Pepe, Francesco, additional, Russo, Gianluca, additional, Pisapia, Pasquale, additional, Gragnano, Gianluca, additional, Aquino, Gabriella, additional, Bessi, Silvia, additional, Buglioni, Simonetta, additional, Bartoccini, Federico, additional, Ferrero, Giuseppina, additional, Bresciani, Michela Anna, additional, Francia di Celle, Paola, additional, Sibona, Francesca, additional, Giusti, Andrea, additional, Movilia, Alessandra, additional, Farioli, Renata Mariella, additional, Santoro, Alessandra, additional, Salemi, Domenico, additional, Scarpino, Stefania, additional, Galafate, Dino, additional, Tommasi, Stefania, additional, Lacalamita, Rosanna, additional, Seminati, Davide, additional, Sajjadi, Elham, additional, Novello, Silvia, additional, Pagni, Fabio, additional, Troncone, Giancarlo, additional, and Malapelle, Umberto, additional
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- 2023
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9. FULLY AUTOMATED REAL-TIME PCR IN MOLECULAR PATHOLOGY: NOVEL APLLICATIONS
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Gragnano, Gianluca
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A deeper knowledge of cancer biology, alongside advances in diagnostic technologies, has led to much more effective therapeutic strategies in Indeed, over the last decade or so, the incessant quest for new genetic alterations capable of predicting patient response to treatments, has given rise to the development of highly sophisticated molecular tests Accordingly, many sequencing platforms can nowadays detect multiple driver mutations simultaneously thanks to fully automated procedures which drastically reduce turnaround time and costs. A case in point is the Idylla™ system (Biocartis NV, Mechelen, Belgium. This system is a fully automated sample-to-result Real-Time PCR with all reagents integrated in a single-use cartridge; it consists of a console which displays results as either "mutation" or "no mutation" when DNA quality is adequate, or as "invalid" when it is. What is most striking about this platform is that the cartridge can run multiple sample types, including solid and liquid biopsies, yielding results within approximately two hours. Another advantage is its ease of use taking up only a couple of minutes of hands-on time to prepare samples and load the cartridge. This fascinating assay has been validated for the identification of many driver genes, both on tissue and on plasma specimens. In particular Idylla™ EGFR, Idylla™ BRAF, Idylla™ KRAS, Idylla™ NRAS-BRAF, and Idylla™ MSI have been validated on formalin fixed paraffin embedded (FFPE) samples; instead, Idylla™ ctKRAS and Idylla™ ctNRAS-BRAF have been validated on plasma samples. Many more are currently in the pipeline. The aim of my PhD project was to develop and validate novel clinical applications of the the Idylla™ system. I divided my thesis into four chapters. The first chapter focuses on the performance of the Idylla™ EGFR Assay on cytological non-small cell lung cancer samples. This test was specifically designed to process formalin-fixed, paraffin-embedded sections without requiring preliminary DNA extraction. On the other hand, my PhD work has demonstrated that this assay can also be used to process archival smears from patients with NSCLC by scraping the stained cellular material directly into the cartridge. The second chapter focuses on the viable application of Idylla™ NRAS-BRAF Assay to cytological thyroid fine-needle aspirates (FNAs) with undetermined morphology. Our research demonstrated that FNA needle rinses can be genotyped by the same cytopathologist who performs the FNA, a procedure that is commonly called rapid on-site molecular evaluation (ROME). The third chapter, instead, discusses the feasibility of using Idylla™ to analyze liquid biopsy specimens. Such application is highly important, given the scant availability of tissue specimens in advanced NSCLC patients. Our laboratory results have indeed demonstrated the efficiency of the Idylla™ ctKRAS Assay in detecting plasma the KRAS p.G12C mutation, a novel target in NSCLC patients. Finally, in the fourth chapter, I explore the analytical and clinical performance of the Idylla™ SARS-CoV-2 test on previously tested SARS-CoV-2 people by conventional RT-PCR based approach in different settings, including initial diagnosis and clinical follow-up. In this regard, I provide substantial evidence that this assay may represent a valid, fast, and highly sensitive and specific RT-PCR test for the identification of SARS-CoV-2 infection.
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- 2021
10. Methods for actionable gene fusion detection in lung cancer: now and in the future
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Pisapia, Pasquale, primary, Pepe, Francesco, additional, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Russo, Gianluca, additional, Gragnano, Gianluca, additional, Conticelli, Floriana, additional, Salatiello, Maria, additional, Luca, Caterina De, additional, Girolami, Ilaria, additional, Eccher, Albino, additional, Iaccarino, Antonino, additional, Bellevicine, Claudio, additional, Vigliar, Elena, additional, Malapelle, Umberto, additional, and Troncone, Giancarlo, additional
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- 2021
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11. Evaluation of Micro Satellite Instability and Mismatch Repair Status in Different Solid Tumors: A Multicenter Analysis in a Real World Setting
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Malapelle, Umberto, primary, Parente, Paola, additional, Pepe, Francesco, additional, De Luca, Caterina, additional, Pisapia, Pasquale, additional, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Gragnano, Gianluca, additional, Russo, Gianluca, additional, Conticelli, Floriana, additional, Bellevicine, Claudio, additional, Vigliar, Elena, additional, Iaccarino, Antonino, additional, Covelli, Claudia, additional, Balistreri, Mariangela, additional, Clemente, Celeste, additional, Perrone, Giovanni, additional, Danza, Angela, additional, Scaramuzzi, Fabio, additional, Fassan, Matteo, additional, Troncone, Giancarlo, additional, and Graziano, Paolo, additional
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- 2021
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12. Liquid Biopsy Analysis in Clinical Practice: Focus on Lung Cancer
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Pisapia, Pasquale, primary, Pepe, Francesco, additional, Iaccarino, Antonino, additional, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Russo, Gianluca, additional, Gragnano, Gianluca, additional, Mosaieby, Elalah, additional, Troncone, Giancarlo, additional, and Malapelle, Umberto, additional
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- 2021
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13. Evaluation of a fully closed real time PCR platform for the detection of SARS-CoV-2 in nasopharyngeal swabs: a pilot study.
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De Luca, Caterina, Gragnano, Gianluca, Conticelli, Floriana, Cennamo, Michele, Pisapia, Pasquale, Terracciano, Daniela, Malapelle, Umberto, Montella, Emma, Triassi, Maria, Troncone, Giancarlo, and Portella, Giuseppe
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COVID-19 ,SARS-CoV-2 ,INTERNET servers - Published
- 2022
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14. PD‐L1 and beyond: Immuno‐oncology in cytopathology
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Iaccarino, Antonino, primary, Salatiello, Maria, additional, Migliatico, Ilaria, additional, De Luca, Caterina, additional, Gragnano, Gianluca, additional, Russo, Maria, additional, Bellevicine, Claudio, additional, Malapelle, Umberto, additional, Troncone, Giancarlo, additional, and Vigliar, Elena, additional
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- 2021
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15. Evaluation of a fully closed real time PCR platform for the detection of SARS-CoV-2 in nasopharyngeal swabs: a pilot study
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De Luca, Caterina, primary, Gragnano, Gianluca, additional, Conticelli, Floriana, additional, Cennamo, Michele, additional, Pisapia, Pasquale, additional, Terracciano, Daniela, additional, Malapelle, Umberto, additional, Montella, Emma, additional, Triassi, Maria, additional, Troncone, Giancarlo, additional, and Portella, Giuseppe, additional
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- 2021
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16. Performance evaluation of a fully closed real-time PCR platform for the detection of KRAS p.G12C mutations in liquid biopsy of patients with non-small cell lung cancer.
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Gragnano, Gianluca, Nacchio, Mariantonia, Sgariglia, Roberta, Conticelli, Floriana, Iaccarino, Antonino, De Luca, Caterina, Troncone, Giancarlo, and Malapelle, Umberto
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NON-small-cell lung carcinoma ,RAS oncogenes ,SMALL cell lung cancer ,PROGRAMMED cell death 1 receptors - Published
- 2022
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17. Performance evaluation of a fully closed real-time PCR platform for the detection of KRAS p.G12C mutations in liquid biopsy of patients with non-small cell lung cancer
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Gragnano, Gianluca, primary, Nacchio, Mariantonia, additional, Sgariglia, Roberta, additional, Conticelli, Floriana, additional, Iaccarino, Antonino, additional, De Luca, Caterina, additional, Troncone, Giancarlo, additional, and Malapelle, Umberto, additional
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- 2021
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18. Next Generation Sequencing in Cytopathology: Focus on Non-Small Cell Lung Cancer
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Pisapia, Pasquale, primary, Pepe, Francesco, additional, Iaccarino, Antonino, additional, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Conticelli, Floriana, additional, Salatiello, Maria, additional, Tufano, Rossella, additional, Russo, Gianluca, additional, Gragnano, Gianluca, additional, Girolami, Ilaria, additional, Eccher, Albino, additional, Malapelle, Umberto, additional, and Troncone, Giancarlo, additional
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- 2021
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19. RNA-Based Assay for Next-Generation Sequencing of Clinically Relevant Gene Fusions in Non-Small Cell Lung Cancer
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De Luca, Caterina, primary, Pepe, Francesco, additional, Iaccarino, Antonino, additional, Pisapia, Pasquale, additional, Righi, Luisella, additional, Listì, Angela, additional, Greco, Lorenza, additional, Gragnano, Gianluca, additional, Campione, Severo, additional, De Dominicis, Gianfranco, additional, Pagni, Fabio, additional, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Tufano, Rossella, additional, Conticelli, Floriana, additional, Vigliar, Elena, additional, Bellevicine, Claudio, additional, Cortinovis, Diego Luigi, additional, Novello, Silvia, additional, Molina-Vila, Miguel Angel, additional, Rosell, Rafael, additional, Troncone, Giancarlo, additional, and Malapelle, Umberto, additional
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- 2021
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20. Tumor mutational burden on cytological samples: A pilot study
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Pepe, Francesco, primary, Pisapia, Pasquale, additional, Gristina, Valerio, additional, Rocco, Danilo, additional, Micheli, Mariacarolina, additional, Micheli, Pietro, additional, Iaccarino, Antonino, additional, Tufano, Rossella, additional, Gragnano, Gianluca, additional, Russo, Gianluca, additional, De Luca, Caterina, additional, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Girolami, Ilaria, additional, Eccher, Albino, additional, Russo, Antonio, additional, Troncone, Giancarlo, additional, and Malapelle, Umberto, additional
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- 2020
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21. Liquid biopsy for BRAF mutations testing in non-small cell lung cancer: a retrospective study
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Iaccarino, Antonino, primary, Pisapia, Pasquale, additional, Pepe, Francesco, additional, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Russo, Gianluca, additional, Gragnano, Gianluca, additional, De Luca, Caterina, additional, Troncone, Giancarlo, additional, and Malapelle, Umberto, additional
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- 2020
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22. BRAF: A Two-Faced Janus
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Pisapia, Pasquale, primary, Pepe, Francesco, additional, Iaccarino, Antonino, additional, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Russo, Gianluca, additional, Gragnano, Gianluca, additional, Malapelle, Umberto, additional, and Troncone, Giancarlo, additional
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- 2020
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23. Concomitant Rare KRAS and BRAF Mutations in Lung Adenocarcinoma: A Case Report
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Iaccarino, Antonino, primary, Pisapia, Pasquale, additional, De Felice, Marco, additional, Pepe, Francesco, additional, Gragnano, Gianluca, additional, De Luca, Caterina, additional, Ianniello, Giovanni, additional, and Malapelle, Umberto, additional
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- 2020
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24. Rapid On‐site Molecular Evaluation in thyroid cytopathology: A same‐day cytological and molecular diagnosis
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De Luca, Caterina, primary, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Pisapia, Pasquale, additional, Migliatico, Ilaria, additional, Clery, Eduardo, additional, Gragnano, Gianluca, additional, Campione, Severo, additional, Vigliar, Elena, additional, Malapelle, Umberto, additional, De Dominicis, Gianfranco, additional, Bellevicine, Claudio, additional, and Troncone, Giancarlo, additional
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- 2020
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25. Tumor mutational burden on cytological samples: A pilot study.
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Pepe, Francesco, Pisapia, Pasquale, Gristina, Valerio, Rocco, Danilo, Micheli, Mariacarolina, Micheli, Pietro, Iaccarino, Antonino, Tufano, Rossella, Gragnano, Gianluca, Russo, Gianluca, De Luca, Caterina, Sgariglia, Roberta, Nacchio, Mariantonia, Girolami, Ilaria, Eccher, Albino, Russo, Antonio, Troncone, Giancarlo, and Malapelle, Umberto
- Abstract
Background: Immune‐checkpoint inhibitors (ICIs) represent an important treatment option for patients who have advanced stage non–small cell lung cancer (NSCLC). Currently, evaluation of the expression level of programmed death‐ligand 1 (PD‐L1) has proven highly successful as a positive predictive biomarker for ICIs. In addition to PD‐L1, other promising predictive biomarkers are emerging, including high tumor mutational burden (TMB‐H). However, measuring TMB‐H remains challenging for several reasons, among which is the difficulty in obtaining adequate tissue material from NSCLC patients. There are no data in the current literature regarding the possibility of adopting cell blocks (CBs) for TMB evaluation; therefore, our goal was to evaluate the feasibility of analyzing TMB on CBs. Methods: For evaluation of differences in run metric parameters, 8 pairs of histological and CB samples from patients with NSCLC were analyzed using the Oncomine Tumor Mutational Load Assay on Ion Torrent S5 GS next‐generation sequencing (NGS) platform. Results: Most CBs (6/8, 75.0%) were successfully analyzed by adopting the broad NGS panel approach. CBs provided results similar to those obtained on histological matched specimens in terms of median total reads (7207048.80 vs 7558817.80), median mapped reads (7075753.83 vs 7513822.00), median read lengths (115.50 vs. 113.00), median percentage of reads on‐target (97.49% vs. 98.45%), median average reads per amplicon (454.67 vs 476.14), and median uniformity of amplicon coverage (83.52% vs 84.13%). Conclusion: In this pilot study, we demonstrated the technical feasibility of assessing TMB on CBs. Immune‐checkpoint inhibitors (ICIs) are an important treatment option for patients who have advanced stage non–small cell lung cancer, and tumor mutational burden (TMB) is a valid biomarker for ICI administration. Cell blocks may be a suitable starting material for TMB analysis. [ABSTRACT FROM AUTHOR]
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- 2021
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26. Is the Idylla EGFR Mutation Assay feasible on archival stained cytological smears? A pilot study
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De Luca, Caterina, primary, Conticelli, Floriana, additional, Leone, Alvaro, additional, Gragnano, Gianluca, additional, Salatiello, Maria, additional, Galasso, Pasqualina, additional, Pisapia, Pasquale, additional, Grillo, Lucia Rosalba, additional, Iaccarino, Antonino, additional, Vigliar, Elena, additional, Bellevicine, Claudio, additional, Malapelle, Umberto, additional, and Troncone, Giancarlo, additional
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- 2019
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27. Performance analysis of SiRe next-generation sequencing panel in diagnostic setting: focus on NSCLC routine samples
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Pepe, Francesco, primary, De Luca, Caterina, additional, Smeraglio, Riccardo, additional, Pisapia, Pasquale, additional, Sgariglia, Roberta, additional, Nacchio, Mariantonia, additional, Russo, Maria, additional, Serra, Nicola, additional, Rocco, Danilo, additional, Battiloro, Ciro, additional, Ambrosio, Francesca, additional, Gragnano, Gianluca, additional, Vigliar, Elena, additional, Bellevicine, Claudio, additional, Troncone, Giancarlo, additional, and Malapelle, Umberto, additional
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- 2018
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28. Liquid biopsy for BRAF mutations testing in non-small cell lung cancer: a retrospective study.
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Iaccarino, Antonino, Pisapia, Pasquale, Pepe, Francesco, Sgariglia, Roberta, Nacchio, Mariantonia, Russo, Gianluca, Gragnano, Gianluca, De Luca, Caterina, Troncone, Giancarlo, and Malapelle, Umberto
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NON-small-cell lung carcinoma ,MOLECULAR pathology ,BRAF genes ,PLATELET-derived growth factor ,SMALL cell lung cancer - Published
- 2022
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29. Idylla assay and next generation sequencing: an integrated EGFR mutational testing algorithm
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De Luca, Caterina, primary, Rappa, Alessandra G, additional, Gragnano, Gianluca, additional, Malapelle, Umberto, additional, Troncone, Giancarlo, additional, and Barberis, Massimo, additional
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- 2018
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30. Liquid biopsy for BRAFmutations testing in non-small cell lung cancer: a retrospective study
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Iaccarino, Antonino, Pisapia, Pasquale, Pepe, Francesco, Sgariglia, Roberta, Nacchio, Mariantonia, Russo, Gianluca, Gragnano, Gianluca, De Luca, Caterina, Troncone, Giancarlo, and Malapelle, Umberto
- Abstract
V-Raf murine sarcoma viral oncogene homolog B (BRAF) gene mutations have recently been approved to select advanced stages non-small cell lung cancer (NSCLC) patients for tyrosine kinase inhibitors treatments. In this setting, liquid biopsy may represent a valuable option for BRAFmutational testing in patients without tissue availability. Here, we reviewed 196 plasma based liquid biopsies analysed by an in-house developed next generation sequencing panel, termed SiRe. On the overall, 6 (3.1%) out of 196 BRAFmutated cases were identified, with an overall median allelic frequency of 3.4%. Exon 15 p.V600E was the most common detected mutation (2/6, 33.3%). Our data highlighted that the SiRe panel is a robust tool for BRAFmutation assessment on circulating tumour DNA. Further investigation is required to develop a diagnostic algorithm to harmonise BRAFtesting on tissue and blood in advanced stages NSCLC patients.
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- 2022
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31. Performance analysis of SiRe next-generation sequencing panel in diagnostic setting: focus on NSCLC routine samples.
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Pepe, Francesco, De Luca, Caterina, Smeraglio, Riccardo, Pisapia, Pasquale, Sgariglia, Roberta, Nacchio, Mariantonia, Russo, Maria, Serra, Nicola, Rocco, Danilo, Battiloro, Ciro, Ambrosio, Francesca, Gragnano, Gianluca, Vigliar, Elena, Bellevicine, Claudio, Troncone, Giancarlo, and Malapelle, Umberto
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NUCLEOTIDE sequencing ,NON-small-cell lung carcinoma ,LUNG cancer ,CANCER diagnosis ,SOMATIC mutation ,EPIDERMAL growth factor receptors ,PROGRESSION-free survival ,CANCER treatment - Published
- 2019
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32. EGFR mutation detection on lung cancer cytological specimens by the novel fully automated PCR-based Idylla EGFR Mutation Assay
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De Luca, Caterina, primary, Gragnano, Gianluca, additional, Pisapia, Pasquale, additional, Vigliar, Elena, additional, Malapelle, Umberto, additional, Bellevicine, Claudio, additional, and Troncone, Giancarlo, additional
- Published
- 2016
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33. Fully automated PCR detection of KRAS mutations on pancreatic endoscopic ultrasound fine needle aspirates.
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De Biase, Dario, primary, de Luca, Caterina, additional, Gragnano, Gianluca, additional, Malapelle, Umberto, additional, Tallini, Giovanni, additional, and Troncone, Giancarlo, additional
- Published
- 2016
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34. EGFR mutation detection on lung cancer cytological specimens by the rapid and fully integrated Idylla molecular diagnostics system.
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de Luca, Caterina, primary, Gragnano, Gianluca, additional, Malapelle, Umberto, additional, and Troncone, Giancarlo, additional
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- 2016
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35. Is the Idylla EGFRMutation Assay feasible on archival stained cytological smears? A pilot study
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De Luca, Caterina, Conticelli, Floriana, Leone, Alvaro, Gragnano, Gianluca, Salatiello, Maria, Galasso, Pasqualina, Pisapia, Pasquale, Grillo, Lucia Rosalba, Iaccarino, Antonino, Vigliar, Elena, Bellevicine, Claudio, Malapelle, Umberto, and Troncone, Giancarlo
- Abstract
AimThe rapid and fully automated Idylla EGFRMutation Assay has been specifically designed to process formalin-fixed, paraffin-embedded sections without requiring preliminary DNA extraction. This study evaluates whether this approach can also process archival smears from patients with non–small cell lung cancer (NSCLC) by scraping the stained cellular material directly into the cartridge.MethodsThe study was divided into two parts. In the first part, we carried out Idylla EGFRMutation Assay on archival stained smears from 39 patients with NSCLC. Among these, 14 cases harboured a mutation in either exon 19 (n=11) or exon 21 (n=3), previously detected on DNA extracts by fragment length and TaqMan assays. In the second part, we evaluated whether de-staining of the smears could reduce background fluorescence.ResultsThe Idylla EGFRMutation Assay confirmed the presence of EGFRmutation in 11 instances (78.6%). However, concordance was higher for exon 19 deletions (10/11) than for exon 21 p.L858R assessments. Raw data showed a high background fluorescence in channel 2, where the EGFRexon 21 p.L858R mutation was detected. This interference, due to dye residues from the original staining, was partially reduced by de-staining the cytological material.ConclusionsOur data, although preliminary, show that the Idylla EGFRMutation Assay can reliably process most archival smears without requiring preliminary DNA extraction. Results may be further improved by de-staining the cellular material before insertion into the cartridge.
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- 2019
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36. EGFR mutation detection on lung cancer cytological specimens by the novel fully automated PCR-based Idylla EGFR Mutation Assay.
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De Luca, Caterina, Gragnano, Gianluca, Pisapia, Pasquale, Vigliar, Elena, Malapelle, Umberto, Bellevicine, Claudio, and Troncone, Giancarlo
- Subjects
LUNG cancer diagnosis ,EPIDERMAL growth factor receptors ,GENETIC mutation ,CYTOLOGY ,POLYMERASE chain reaction - Published
- 2017
- Full Text
- View/download PDF
37. Fully automated PCR detection of KRAS mutations on pancreatic endoscopic ultrasound fine-needle aspirates.
- Author
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de Biase, Dario, de Luca, Caterina, Gragnano, Gianluca, Visani, Michela, Bellevicine, Claudio, Malapelle, Umberto, Tallini, Giovanni, and Troncone, Giancarlo
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PANCREATIC cancer diagnosis ,GENETIC mutation ,ENDOSCOPY ,POLYMERASE chain reaction ,NUCLEOTIDE sequencing - Published
- 2016
- Full Text
- View/download PDF
38. EGFRmutation detection on lung cancer cytological specimens by the novel fully automated PCR-based Idylla EGFRMutation Assay
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De Luca, Caterina, Gragnano, Gianluca, Pisapia, Pasquale, Vigliar, Elena, Malapelle, Umberto, Bellevicine, Claudio, and Troncone, Giancarlo
- Abstract
AimsIn everyday practice, epidermal growth factor receptor (EGFR) testing is centralised in referral laboratories that receive paucicellular cytological specimens. Ideally, EGFRtesting should be carried out in the centre where the patient is diagnosed such that the most cellular slide can be selected from in-house collected cytological material. However, available techniques are little standardised and difficult to be implemented in settings with little expertise in molecular testing. The Idylla EGFRprototype assay is a rapid and fully automated test which may easily be adopted by a wider number of pathological centres. This study assessed whether an Idylla EGFRprototype assay can be reliably applied to cytological lung cancer specimens.MethodsThe limit of detection (LOD) of the Idylla EGFRprototype assay was assessed by cell line dilution studies. A total of 10 ng was directly placed inside an Idylla EGFRprototype assay cartridge. Idylla results were compared with fragment length (exon 19 del) and Taqman assays.ResultsThe Idylla EGFRprototype assay showed an LOD of 1% mutant allele and yielded valid results in 74/76 (97.3%) samples, detecting all the mutant cases (n=32) identified by standard techniques; in addition, Idylla detected two low abundance EGFRexon 19 deletions and two G719X exon 18 point mutations, not covered by our standard reference method.ConclusionsIdylla EGFRprototype assay is sensitive on extracted DNA and can reliably be applied on cytological samples, enabling implementation of EGFRtesting even in less experienced diagnostic units.
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- 2017
- Full Text
- View/download PDF
39. Liquid Biopsy Analysis in Clinical Practice: Focus on Lung Cancer
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Francesco Pepe, Antonino Iaccarino, Roberta Sgariglia, Gianluca Russo, G. Troncone, Umberto Malapelle, Mariantonia Nacchio, Elalah Mosaieby, Pasquale Pisapia, Gianluca Gragnano, Pisapia, Pasquale, Pepe, Francesco, Iaccarino, Antonino, Sgariglia, Roberta, Nacchio, Mariantonia, Russo, Gianluca, Gragnano, Gianluca, Mosaieby, Elalah, Troncone, Giancarlo, and Malapelle, Umberto
- Subjects
0301 basic medicine ,medicine.medical_specialty ,EGFR ,Molecular oncology ,03 medical and health sciences ,0302 clinical medicine ,molecular oncology ,medicine ,Pathology ,RB1-214 ,Liquid biopsy ,Intensive care medicine ,Lung cancer ,predictive molecular pathology ,predictive biomarker ,business.industry ,Advanced stage ,Treatment options ,ctDNA ,medicine.disease ,Clinical Practice ,030104 developmental biology ,030220 oncology & carcinogenesis ,Sample collection ,Non small cell ,business - Abstract
Lung cancer is the leading cause of cancer death worldwide. Despite the emergence of highly effective targeted therapies, up to 30% of advanced stage non-small cell lung cancer (NSCLC) patients do not undergo tissue molecular testing because of scarce tissue availability. Liquid biopsy, on the other hand, offers these patients a valuable opportunity to receive the best treatment options in a timely manner. Indeed, besides being much faster and less invasive than conventional tissue-based analysis, it can also yield specific information about the genetic make-up and evolution of patients’ tumors. However, several issues, including lack of standardized protocols for sample collection, processing, and interpretation, still need to be addressed before liquid biopsy can be fully incorporated into routine oncology practice. Here, we reviewed the most important challenges hindering the implementation of liquid biopsy in oncology practice, as well as the great advantages of this approach for the treatment of NSCLC patients.
- Published
- 2021
40. An Italian Multicenter Perspective Harmonization Trial for the Assessment of MET Exon 14 Skipping Mutations in Standard Reference Samples
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Paolo Bironzo, Francesco Pepe, Gianluca Russo, Pasquale Pisapia, Gianluca Gragnano, Gabriella Aquino, Silvia Bessi, Simonetta Buglioni, Federico Bartoccini, Giuseppina Ferrero, Michela Anna Bresciani, Paola Francia di Celle, Francesca Sibona, Andrea Giusti, Alessandra Movilia, Renata Mariella Farioli, Alessandra Santoro, Domenico Salemi, Stefania Scarpino, Dino Galafate, Stefania Tommasi, Rosanna Lacalamita, Davide Seminati, Elham Sajjadi, Silvia Novello, Fabio Pagni, Giancarlo Troncone, Umberto Malapelle, Bironzo, P, Pepe, F, Russo, G, Pisapia, P, Gragnano, G, Aquino, G, Bessi, S, Buglioni, S, Bartoccini, F, Ferrero, G, Bresciani, M, Francia di Celle, P, Sibona, F, Giusti, A, Movilia, A, Farioli, R, Santoro, A, Salemi, D, Scarpino, S, Galafate, D, Tommasi, S, Lacalamita, R, Seminati, D, Sajjadi, E, Novello, S, Pagni, F, Troncone, G, Malapelle, U, Bironzo, Paolo, Pepe, Francesco, Russo, Gianluca, Pisapia, Pasquale, Gragnano, Gianluca, Aquino, Gabriella, Bessi, Silvia, Buglioni, Simonetta, Bartoccini, Federico, Ferrero, Giuseppina, Anna Bresciani, Michela, Francia di Celle, Paola, Sibona, Francesca, Giusti, Andrea, Movilia, Alessandra, Mariella Farioli, Renata, Santoro, Alessandra, Salemi, Domenico, Scarpino, Stefania, Galafate, Dino, Tommasi, Stefania, Lacalamita, Rosanna, Seminati, Davide, Sajjadi, Elham, Novello, Silvia, Pagni, Fabio, Troncone, Giancarlo, and Malapelle, Umberto
- Subjects
Clinical Biochemistry ,MET ,NSCLC ,molecular test - Abstract
Lung cancer remains the leading cause of cancer deaths worldwide. International societies have promoted the molecular analysis of MET proto-oncogene, receptor tyrosine kinase (MET) exon 14 skipping for the clinical stratification of non-small cell lung cancer (NSCLC) patients. Different technical approaches are available to detect MET exon 14 skipping in routine practice. Here, the technical performance and reproducibility of testing strategies for MET exon 14 skipping carried out in various centers were evaluated. In this retrospective study, each institution received a set (n = 10) of a customized artificial formalin-fixed paraffin-embedded (FFPE) cell line (Custom METex14 skipping FFPE block) that harbored the MET exon 14 skipping mutation (Seracare Life Sciences, Milford, MA, USA), which was previously validated by the Predictive Molecular Pathology Laboratory at the University of Naples Federico II. Each participating institution managed the reference slides according to their internal routine workflow. MET exon 14 skipping was successfully detected by all participating institutions. Molecular analysis highlighted a median Cq cut off of 29.3 (ranging from 27.1 to 30.7) and 2514 (ranging from 160 to 7526) read counts for real-time polymerase chain reaction (RT-PCR) and NGS-based analyses, respectively. Artificial reference slides were a valid tool to harmonize technical workflows in the evaluation of MET exon 14 skipping molecular alterations in routine practice.
- Published
- 2023
41. Concomitant Rare KRAS and BRAF Mutations in Lung Adenocarcinoma: A Case Report
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Giovanni Pietro Ianniello, Marco De Felice, Umberto Malapelle, Pasquale Pisapia, Gianluca Gragnano, Antonino Iaccarino, Francesco Pepe, Caterina De Luca, Iaccarino, Antonino, Pisapia, Pasquale, De Felice, Marco, Pepe, Francesco, Gragnano, Gianluca, De Luca, Caterina, Ianniello, Giovanni, and Malapelle, Umberto
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0301 basic medicine ,business.industry ,Pleural effusion ,Point mutation ,medicine.disease ,medicine.disease_cause ,digestive system diseases ,03 medical and health sciences ,Exon ,030104 developmental biology ,0302 clinical medicine ,Real-time polymerase chain reaction ,030220 oncology & carcinogenesis ,medicine ,Cancer research ,ROS1 ,Adenocarcinoma ,KRAS ,business ,Lung cancer - Abstract
In July 2020, an active smoker, 63-year old man was admitted to the oncology unit of A.O.R.N. Sant’Anna e San Sebastiano (Caserta, Italy). Chest radiology highlighted right pleural effusion. Total-body CT scanning revealed a solid lesion with lobulated contours in the apical segment of the upper right lobe. The patient’s oncologist requested a molecular assessment of EGFR, ALK, ROS1, BRAF, and KRAS, as well as an evaluation of PD-L1 expression level. To this end, we carried out NGS analysis, on DNA extracted from cytospins, by adopting a custom-designed NGS panel (SiRe®). Overall, no actionable mutations in the tested genes were identified. Conversely, concomitant BRAF exon 11 p.G469A and a KRAS exon 4 p.A146T mutations were detected. Owing to the limited data on the presence of KRAS exon 4 p.A146T point mutation in lung adenocarcinoma patients, a further molecular confirmatory analysis was carried out with a dedicated KRAS cartridge on a fully automated real time polymerase chain reaction. When DNA was extracted from the TTF-1 positive tumor cell slide, the same KRAS alteration was observed. Unfortunately, the patient died in August 2020 before having the chance to start any type of treatment.
- Published
- 2020
42. Performance analysis of SiRe next-generation sequencing panel in diagnostic setting: focus on NSCLC routine samples
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Elena Vigliar, Riccardo Smeraglio, Roberta Sgariglia, Danilo Rocco, Nicola Serra, Umberto Malapelle, Mariantonia Nacchio, Maria Giovanna Russo, Giancarlo Troncone, Claudio Bellevicine, Francesco Pepe, Gianluca Gragnano, Ciro Battiloro, Caterina De Luca, Francesca Ambrosio, Pasquale Pisapia, Pepe, Francesco, De Luca, Caterina, Smeraglio, Riccardo, Pisapia, Pasquale, Sgariglia, Roberta, Nacchio, Mariantonia, Russo, Maria, Serra, Nicola, Rocco, Danilo, Battiloro, Ciro, Ambrosio, Francesca, Gragnano, Gianluca, Vigliar, Elena, Bellevicine, Claudio, Troncone, Giancarlo, and Malapelle, Umberto
- Subjects
Male ,0301 basic medicine ,Oncology ,Neuroblastoma RAS viral oncogene homolog ,medicine.medical_specialty ,Lung Neoplasms ,PDGFRA ,NSCLC ,medicine.disease_cause ,DNA sequencing ,Pathology and Forensic Medicine ,03 medical and health sciences ,0302 clinical medicine ,Carcinoma, Non-Small-Cell Lung ,Internal medicine ,Biomarkers, Tumor ,medicine ,Humans ,predictive molecular pathology ,Lung cancer ,business.industry ,Molecular pathology ,Sire ,High-Throughput Nucleotide Sequencing ,Sequence Analysis, DNA ,General Medicine ,Middle Aged ,Amplicon ,medicine.disease ,Lung Neoplasm ,030104 developmental biology ,NGS ,030220 oncology & carcinogenesis ,Mutation ,Female ,KRAS ,business ,Human - Abstract
AimsFollowing the development for liquid biopsies of the SiRe next-generation sequencing (NGS) panel that covers 568 clinical relevant mutations in EGFR, KRAS, NRAS, BRAF, cKIT and PDGFRa genes, in this current study, we apply this small NGS panel on tissue samples of lung cancer.MethodsA total of 322 specimens were prospectively tested. Technical parameters were analysed on both cytological and histological samples. In a subset of 75 samples, the EGFR SiRe results were compared with those generated by the European Community (CE)–IVD EGFR assay on Idylla platform. Clinical outcomes of 11 patients treated, on the basis of SiRe results, were also evaluated.ResultsOnly 28 (8.7%) specimens failed to produce a library; out of the 294 remaining samples, a total of 168 somatic mutations were found. In nearly all instances (74/75–99%), the EGFR SiRe results were confirmed by Idylla. In general, SiRe analytical parameters were excellent. However, histological and cytological specimens differed in relation to average reads for sample, mean number of mapped reads, median read length and average reads for amplicon. Treatment outcome evaluation in 11 patients showed a partial response in 82 % (9/11) patients with a median progression-free survival of 340 days.ConclusionsThe small gene panel SiRe is a clinically relevant tool useful to widespread the adoption of NGS in predictive molecular pathology laboratories.
- Published
- 2018
43. Next Generation Sequencing in Cytopathology: Focus on Non-Small Cell Lung Cancer
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Giancarlo Troncone, Roberta Sgariglia, Francesco Pepe, Pasquale Pisapia, Mariantonia Nacchio, Ilaria Girolami, Umberto Malapelle, Antonino Iaccarino, Gianluca Gragnano, Albino Eccher, Floriana Conticelli, Maria Salatiello, Rossella Tufano, Gianluca Russo, Pisapia, Pasquale, Pepe, Francesco, Iaccarino, Antonino, Sgariglia, Roberta, Nacchio, Mariantonia, Conticelli, Floriana, Salatiello, Maria, Tufano, Rossella, Russo, Gianluca, Gragnano, Gianluca, Girolami, Ilaria, Eccher, Albino, Malapelle, Umberto, and Troncone, Giancarlo
- Subjects
lcsh:R5-920 ,Focus (geometry) ,medicine.diagnostic_test ,Molecular pathology ,Mini Review ,fine needle aspiration ,smear ,General Medicine ,Computational biology ,Biology ,NSCLC ,cell block ,medicine.disease ,DNA sequencing ,Fine-needle aspiration ,cytopathology ,liquid based cytology ,molecular cytopathology ,Cytopathology ,Liquid-based cytology ,medicine ,Medicine ,Non small cell ,lcsh:Medicine (General) ,Lung cancer - Abstract
Molecular cytopathology is a rapidly evolving field embracing both conventional microscopy and molecular pathology. Its growing popularity stems from the fact that in many types of advanced cancers, including non small cell lung cancer (NSCLC), cytological samples often constitute the only available specimens for morphomolecular analysis. Indeed, non formalin fixed and paraffin embedded (FFPE) cytological samples feature a higher quality of extracted nucleic acids than histological specimens. However, because of the growing complexity of molecular testing, several efforts should be made to validate the analytical performance of the wide array of currently available molecular technologies, including next generation sequencing (NGS). This technology has the terrific advantage of allowing simultaneous detection of scores of predictive biomarkers even in low-input DNA/RNA specimens. Here, we briefly review the role of the modern cytopathologist in the morphomolecular diagnosing of advanced stage NSCLC and the adoption of NGS in conventional cytopreparations (cell blocks, direct smears, and liquid-based cytology) and supernatants.
- Published
- 2021
44. Evaluation of a fully closed real time PCR platform for the detection of SARS-CoV-2 in nasopharyngeal swabs: a pilot study
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Daniela Terracciano, Umberto Malapelle, Giancarlo Troncone, Emma Montella, Michele Cennamo, Caterina De Luca, Pasquale Pisapia, Gianluca Gragnano, Maria Triassi, Giuseppe Portella, Floriana Conticelli, De Luca, Caterina, Gragnano, Gianluca, Conticelli, Floriana, Cennamo, Michele, Pisapia, Pasquale, Terracciano, Daniela, Malapelle, Umberto, Montella, Emma, Triassi, Maria, Troncone, Giancarlo, and Portella, Giuseppe
- Subjects
0301 basic medicine ,medicine.medical_specialty ,Coronavirus disease 2019 (COVID-19) ,Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ,Pilot Projects ,Real-Time Polymerase Chain Reaction ,Sensitivity and Specificity ,Pathology and Forensic Medicine ,03 medical and health sciences ,0302 clinical medicine ,Internal medicine ,Nasopharynx ,medicine ,Humans ,Close contact ,Original Research ,Cycle threshold ,business.industry ,SARS-CoV-2 ,Clinical performance ,COVID-19 ,General Medicine ,Gold standard (test) ,infection control ,Highly sensitive ,030104 developmental biology ,Real-time polymerase chain reaction ,PCR ,030220 oncology & carcinogenesis ,business - Abstract
AimsTo date, reverse transcriptase PCR (RT-PCR) on nasopharyngeal swabs is the ‘gold standard’ approach for the diagnosis of COVID-19. The need to develop easy to use, rapid, robust and with minimal hands-on time approaches are warranted. In this setting, the Idylla SARS-CoV-2 Test may be a valuable option. The aim of our study is to evaluate the analytical and clinical performance of this assay on previously tested SARS-CoV-2 people by conventional RT-PCR based approach in different settings, including initial diagnosis and clinical follow-up.MethodsTo evaluate the sensitivity and specificity of the Idylla SARS-CoV-2 Test, we retrieved 55 nasopharyngeal swabs, previously analysed by a fully validated assay, from symptomatic patients or from people who have been in close contact with COVID-19 positive cases. Discordant or high discrepant cases were further analysed by a third technique. In addition, a second subset of 14 nasopharyngeal swab samples with uncertain results (cycle threshold between 37 and 40), by using the fully validated assay, from patients with viral infection beyond day 21, were retrieved.ResultsOverall, Idylla showed a sensitivity of 93.9% and a specificity of 100.0%. In addition, in the additional 14 nasopharyngeal swab samples, only five (35.7%) featured a positive result by the Idylla SARS-CoV-2 Test.ConclusionsWe demonstrated that the Idylla SARS-CoV-2 Test may represent a valid, fast, highly sensitive and specific RT-PCR test for the identification of SARS-CoV-2 infection.
- Published
- 2021
45. Evaluation of Micro Satellite Instability and Mismatch Repair Status in Different Solid Tumors: A Multicenter Analysis in a Real World Setting
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Roberta Sgariglia, Antonino Iaccarino, Caterina De Luca, Pasquale Pisapia, Mariangela Balistreri, Giancarlo Troncone, Angela Danza, Celeste Clemente, Claudio Bellevicine, Umberto Malapelle, Paola Parente, Francesco Pepe, Paolo Graziano, Giovanni Perrone, Fabio Scaramuzzi, Claudia Covelli, Elena Vigliar, Gianluca Russo, Gianluca Gragnano, Mariantonia Nacchio, Matteo Fassan, Floriana Conticelli, Malapelle, Umberto, Parente, Paola, Pepe, Francesco, De Luca, Caterina, Pisapia, Pasquale, Sgariglia, Roberta, Nacchio, Mariantonia, Gragnano, Gianluca, Russo, Gianluca, Conticelli, Floriana, Bellevicine, Claudio, Vigliar, Elena, Iaccarino, Antonino, Covelli, Claudia, Balistreri, Mariangela, Clemente, Celeste, Perrone, Giovanni, Danza, Angela, Scaramuzzi, Fabio, Fassan, Matteo, Troncone, Giancarlo, and Graziano, Paolo
- Subjects
Male ,0301 basic medicine ,Oncology ,Colorectal cancer ,microfluidic ,Digestive System Neoplasms ,DNA Mismatch Repair ,Polymerase Chain Reaction ,0302 clinical medicine ,Biology (General) ,immune checkpoint inhibitors (ICIs) ,Ovarian Neoplasms ,Sanger sequencing ,Fully automated RT-PCR ,IHC ,Immune checkpoint inhibitors (ICIs) ,Immunotherapy ,Microfluidic ,MMR ,MSI ,Predictive molecular pathology ,General Medicine ,Microfluidic Analytical Techniques ,Immunohistochemistry ,Italy ,Molecular Diagnostic Techniques ,030220 oncology & carcinogenesis ,symbols ,Female ,Microsatellite Instability ,DNA mismatch repair ,immunotherapy ,medicine.medical_specialty ,Genital Neoplasms, Female ,QH301-705.5 ,Concordance ,Article ,03 medical and health sciences ,symbols.namesake ,Predictive Value of Tests ,Stomach Neoplasms ,Internal medicine ,Biomarkers, Tumor ,medicine ,Humans ,predictive molecular pathology ,neoplasms ,Retrospective Studies ,business.industry ,Advanced stage ,Prostatic Neoplasms ,Reproducibility of Results ,Microsatellite instability ,fully automated RT-PCR ,medicine.disease ,digestive system diseases ,Endometrial Neoplasms ,Pancreatic Neoplasms ,DNA Repair Enzymes ,030104 developmental biology ,Microsatellite Stable ,business - Abstract
Immune-checkpoint inhibitors (ICIs) play a key role in the treatment of advanced stage colorectal cancer (CRC) patients featuring a deficient DNA mismatch repair (dMMR) system or a high microsatellite instability (MSI-H) profile. However, beyond the established role in CRC patients, ICIs have highly proven efficacy in other solid tumors featuring MSI-H/dMMR status represented by endometrial, gastric, ovarian, prostatic, and pancreatic carcinomas (EC, GC, OC, PrC, and PaC). Our aim was to compare the concordance rates among the Idylla™ MSI test, TapeStation 4200, and immunohistochemical (IHC) analysis in assessing MSI-H/dMMR status in EC, GC, OC, PrC, and PaC patients. The Sanger sequencing-based Titano MSI test was used in discordant cases. One hundred and eighty-five cases (n = 40 PrC, n = 39 GC, n = 38 OC, n = 35 PaC, and n = 33 EC) were retrospectively selected. MMR protein expression was evaluated by IHC. After DNA quality and quantity evaluations, the IdyllaTM and TapeStation 4200 platforms were adopted for the evaluation of MSI status. Remarkably, compared to IHC, the Idylla™ platform achieved a global concordance rate of 94.5% (154/163) for the microsatellite stable (MSS)/proficient MMR (pMMR) cases and 77.3% (17/22) for the MSI-H/dMMR cases. Similarly, a global concordance rate of 91.4% (149/163) and 68.2% (15/22) for MSS/pMMR and MSI-H/dMMR cases was also identified between IHC and the TapeStation 4200 microfluidic system. In addition, a global concordance of 93.1% (148/159) and 69.2% (18/26) for MSS/pMMR and MSI-H/dMMR cases was observed between the Idylla™ and TapeStation 4200 platforms. Discordant cases were analyzed using the Titano MSI kit. Overall, our data pinpointed a central role for molecular techniques in the diagnostic evaluation of dMMR/MSI-H status not only in CRC patients but also in other types of solid tumors.
- Published
- 2021
46. Next Generation Sequencing for liquid biopsy based testing in Non-Small Cell Lung Cancer in 2021
- Author
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Francesco Pepe, Gianluca Russo, Antonino Iaccarino, Gianluca Gragnano, José Luis Costa, Alessandro Russo, Luca Quagliata, María José Serrano, Christian Rolfo, Umberto Malapelle, Diego de Miguel-Pérez, Valeria Denninghoff, Pasquale Pisapia, Pisapia, Pasquale, Costa, José Lui, Pepe, Francesco, Russo, Gianluca, Gragnano, Gianluca, Russo, Alessandro, Iaccarino, Antonino, de Miguel-Perez, Diego, Serrano, Maria Josè, Denninghoff, Valeria, Quagliata, Luca, Rolfo, Christian, and Malapelle, Umberto
- Subjects
0301 basic medicine ,Oncology ,medicine.medical_specialty ,Lung Neoplasms ,Context (language use) ,NSCLC ,DNA sequencing ,03 medical and health sciences ,0302 clinical medicine ,Internal medicine ,Carcinoma, Non-Small-Cell Lung ,Biomarkers, Tumor ,Medicine ,Humans ,Liquid biopsy ,Lung cancer ,early detection ,liquid biopsy ,business.industry ,Molecular pathology ,screening ,Advanced stage ,Cancer ,High-Throughput Nucleotide Sequencing ,Hematology ,medicine.disease ,030104 developmental biology ,030220 oncology & carcinogenesis ,NGS ,Non small cell ,business - Abstract
Lung cancer is the leading cause of cancer death worldwide, with non-small cell lung cancer (NSCLC) representing its most commonly diagnosed sub-type. Despite the significant improvements in lung cancer biomarkers knowledge, accompanied by substantial technological advances in molecular tumor profiling, a considerable fraction (up to 30 %) of advanced NSCLC patient presents with major testing challenges or tissue unavailability for molecular analysis. In this context, liquid biopsy is on the rise, currently gaining considerable interest within the molecular pathology and oncology community. Molecular profiling of liquid biopsy specimens using next generation molecular biology methodologies is a rapidly evolving field with promising applications not exclusively limited to advanced stages but also more recently expanding to early stages cancer patients. Here, we offer an overview of some of the most consolidated and emerging applications of next generation sequencing technologies for liquid biopsy testing in NSCLC.
- Published
- 2021
47. Methods for actionable gene fusion detection in lung cancer: now and in the future
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Mariantonia Nacchio, Ilaria Girolami, Roberta Sgariglia, Umberto Malapelle, Elena Vigliar, Floriana Conticelli, Antonino Iaccarino, Gianluca Gragnano, Claudio Bellevicine, Pasquale Pisapia, Albino Eccher, Giancarlo Troncone, Gianluca Russo, Francesco Pepe, Maria Salatiello, Caterina De Luca, Pisapia, Pasquale, Pepe, Francesco, Sgariglia, Roberta, Nacchio, Mariantonia, Russo, Gianluca, Gragnano, Gianluca, Conticelli, Floriana, Salatiello, Maria, Luca, Caterina De, Girolami, Ilaria, Eccher, Albino, Iaccarino, Antonino, Bellevicine, Claudio, Vigliar, Elena, Malapelle, Umberto, and Troncone, Giancarlo
- Subjects
Lung Neoplasms ,Computational biology ,Medical Oncology ,Fusion gene ,Carcinoma, Non-Small-Cell Lung ,Genetics ,medicine ,ROS1 ,Humans ,Routine clinical practice ,Lung cancer ,Pharmacology ,medicine.diagnostic_test ,business.industry ,High-Throughput Nucleotide Sequencing ,medicine.disease ,Clinical trial ,ALK ,NGS ,nCounter ,Molecular Medicine ,Tissue material ,Treatment decision making ,Gene Fusion ,NTRK ,RET ,business ,Fluorescence in situ hybridization - Abstract
Although gene fusions occur rarely in non-small-cell lung cancer (NSCLC) patients, they represent a relevant target in treatment decision algorithms. To date, immunohistochemistry and fluorescence in situ hybridization are the two principal methods used in clinical trials. However, using these methods in routine clinical practice is often impractical and time consuming because they can only analyze single genes and the quantity of tissue material is often insufficient. Thus, novel technologies, able to test multiple genes in a single run with minimal sample input, are being under investigation. Here, we discuss the utility of next-generation sequencing and nCounter technologies in detecting simultaneous gene fusions in NSCLC patients.
- Published
- 2021
48. RNA-Based Assay for Next-Generation Sequencing of Clinically Relevant Gene Fusions in Non-Small Cell Lung Cancer
- Author
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Fabio Pagni, Caterina De Luca, Diego Cortinovis, Silvia Novello, Luisella Righi, Roberta Sgariglia, Miguel Angel Molina-Vila, Lorenza Greco, Floriana Conticelli, Gianfranco De Dominicis, Pasquale Pisapia, Antonino Iaccarino, Claudio Bellevicine, Angela Listì, Rossella Tufano, Francesco Pepe, Elena Vigliar, Giancarlo Troncone, Umberto Malapelle, Rafael Rosell, Gianluca Gragnano, Severo Campione, Mariantonia Nacchio, De Luca, C, Pepe, F, Iaccarino, A, Pisapia, P, Righi, L, Listì, A, Greco, L, Gragnano, G, Campione, S, De Dominicis, G, Pagni, F, Sgariglia, R, Nacchio, M, Tufano, R, Conticelli, F, Vigliar, E, Bellevicine, C, Cortinovis, D, Novello, S, Molina-Vila, M, Rosell, R, Troncone, G, Malapelle, U, De Luca, Caterina, Pepe, Francesco, Iaccarino, Antonino, Pisapia, Pasquale, Righi, Luisella, Listì, Angela, Greco, Lorenza, Gragnano, Gianluca, Campione, Severo, De Dominicis, Gianfranco, Pagni, Fabio, Sgariglia, Roberta, Nacchio, Mariantonia, Tufano, Rossella, Conticelli, Floriana, Vigliar, Elena, Bellevicine, Claudio, Cortinovis, Diego Luigi, Novello, Silvia, Molina-Vila, Miguel Angel, Rosell, Rafael, Troncone, Giancarlo, and Malapelle, Umberto
- Subjects
0301 basic medicine ,Cancer Research ,Computational biology ,NSCLC ,gene fusions ,next generation sequencing ,predictive molecular pathology ,targeted therapy ,Biology ,lcsh:RC254-282 ,Article ,DNA sequencing ,law.invention ,03 medical and health sciences ,0302 clinical medicine ,law ,Complementary DNA ,medicine ,Lung cancer ,Gene ,Polymerase chain reaction ,gene fusion ,RNA ,medicine.disease ,lcsh:Neoplasms. Tumors. Oncology. Including cancer and carcinogens ,030104 developmental biology ,Oncology ,NGS ,030220 oncology & carcinogenesis ,RNA splicing ,Adenocarcinoma - Abstract
Gene fusions represent novel predictive biomarkers for advanced non-small cell lung cancer (NSCLC). In this study, we validated a narrow NGS gene panel able to cover therapeutically-relevant gene fusions and splicing events in advanced-stage NSCLC patients. To this aim, we first assessed minimal complementary DNA (cDNA) input and the limit of detection (LoD) in different cell lines. Then, to evaluate the feasibility of applying our panel to routine clinical samples, we retrospectively selected archived lung adenocarcinoma histological and cytological (cell blocks) samples. Overall, our SiRe RNA fusion panel was able to detect all fusions and a splicing event harbored in a RNA pool diluted up to 2 ng/µ, L. It also successfully analyzed 46 (95.8%) out of 48 samples. Among these, 43 (93.5%) out of 46 samples reproduced the same results as those obtained with conventional techniques. Intriguingly, the three discordant results were confirmed by a CE-IVD automated real-time polymerase chain reaction (RT-PCR) analysis (Easy PGX platform, Diatech Pharmacogenetics, Jesi, Italy). Based on these findings, we conclude that our new SiRe RNA fusion panel is a valid and robust tool for the detection of clinically relevant gene fusions and splicing events in advanced NSCLC.
- Published
- 2021
49. Tumor mutational burden on cytological samples: A pilot study
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Roberta Sgariglia, Giancarlo Troncone, Francesco Pepe, Antonino Iaccarino, Albino Eccher, Mariantonia Nacchio, Rossella Tufano, Mariacarolina Micheli, Pasquale Pisapia, Danilo Rocco, Ilaria Girolami, Umberto Malapelle, Gianluca Russo, Gianluca Gragnano, Pietro Micheli, Antonio Russo, Caterina De Luca, Valerio Gristina, Pepe F., Pisapia P., Gristina V., Rocco D., Micheli M., Micheli P., Iaccarino A., Tufano R., Gragnano G., Russo G., De Luca C., Sgariglia R., Nacchio M., Girolami I., Eccher A., Russo A., Troncone G., Malapelle U., Pepe, Francesco, Pisapia, Pasquale, Gristina, Valerio, Rocco, Danilo, Micheli, Mariacarolina, Micheli, Pietro, Iaccarino, Antonino, Tufano, Rossella, Gragnano, Gianluca, Russo, Gianluca, De Luca, Caterina, Sgariglia, Roberta, Nacchio, Mariantonia, Girolami, Ilaria, Eccher, Albino, Russo, Antonio, Troncone, Giancarlo, and Malapelle, Umberto
- Subjects
Oncology ,Male ,Cancer Research ,medicine.medical_specialty ,Lung Neoplasms ,Cytological Techniques ,DNA Mutational Analysis ,030209 endocrinology & metabolism ,Pilot Projects ,03 medical and health sciences ,0302 clinical medicine ,Internal medicine ,Carcinoma, Non-Small-Cell Lung ,medicine ,Biomarkers, Tumor ,Humans ,Lung cancer ,Predictive biomarker ,Aged ,Retrospective Studies ,next generation sequencing ,TMB ,business.industry ,Advanced stage ,Treatment options ,High-Throughput Nucleotide Sequencing ,Ion semiconductor sequencing ,Amplicon ,medicine.disease ,Prognosis ,lung cancer ,030220 oncology & carcinogenesis ,Mutation ,cytology ,Tissue material ,Female ,immunotherapy ,Non small cell ,business ,Follow-Up Studies - Abstract
Background Immune-checkpoint inhibitors (ICIs) represent an important treatment option for patients who have advanced stage non-small cell lung cancer (NSCLC). Currently, evaluation of the expression level of programmed death-ligand 1 (PD-L1) has proven highly successful as a positive predictive biomarker for ICIs. In addition to PD-L1, other promising predictive biomarkers are emerging, including high tumor mutational burden (TMB-H). However, measuring TMB-H remains challenging for several reasons, among which is the difficulty in obtaining adequate tissue material from NSCLC patients. There are no data in the current literature regarding the possibility of adopting cell blocks (CBs) for TMB evaluation; therefore, our goal was to evaluate the feasibility of analyzing TMB on CBs. Methods For evaluation of differences in run metric parameters, 8 pairs of histological and CB samples from patients with NSCLC were analyzed using the Oncomine Tumor Mutational Load Assay on Ion Torrent S5 GS next-generation sequencing (NGS) platform. Results Most CBs (6/8, 75.0%) were successfully analyzed by adopting the broad NGS panel approach. CBs provided results similar to those obtained on histological matched specimens in terms of median total reads (7207048.80 vs 7558817.80), median mapped reads (7075753.83 vs 7513822.00), median read lengths (115.50 vs. 113.00), median percentage of reads on-target (97.49% vs. 98.45%), median average reads per amplicon (454.67 vs 476.14), and median uniformity of amplicon coverage (83.52% vs 84.13%). Conclusion In this pilot study, we demonstrated the technical feasibility of assessing TMB on CBs.
- Published
- 2020
50. EGFR mutation detection on lung cancer cytological specimens by the novel fully automated PCR-based Idylla EGFR Mutation Assay
- Author
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Gianluca Gragnano, Umberto Malapelle, Elena Vigliar, Claudio Bellevicine, Giancarlo Troncone, Caterina De Luca, Pasquale Pisapia, De Luca, Caterina, Gragnano, Gianluca, Pisapia, Pasquale, Vigliar, Elena, Malapelle, Umberto, Bellevicine, Claudio, and Troncone, Giancarlo
- Subjects
0301 basic medicine ,EGFR ,Pathology and Forensic Medicine ,law.invention ,CYTOLOGY ,03 medical and health sciences ,Exon ,0302 clinical medicine ,law ,TaqMan ,medicine ,Epidermal growth factor receptor ,Lung cancer ,MOLECULAR PATHOLOGY ,Polymerase chain reaction ,biology ,Molecular pathology ,Point mutation ,LUNG CANCER ,General Medicine ,medicine.disease ,Molecular biology ,030104 developmental biology ,Fully automated ,030220 oncology & carcinogenesis ,biology.protein - Abstract
AimsIn everyday practice, epidermal growth factor receptor (EGFR) testing is centralised in referral laboratories that receive paucicellular cytological specimens. Ideally, EGFR testing should be carried out in the centre where the patient is diagnosed such that the most cellular slide can be selected from in-house collected cytological material. However, available techniques are little standardised and difficult to be implemented in settings with little expertise in molecular testing. The Idylla EGFR prototype assay is a rapid and fully automated test which may easily be adopted by a wider number of pathological centres. This study assessed whether an Idylla EGFR prototype assay can be reliably applied to cytological lung cancer specimens.MethodsThe limit of detection (LOD) of the Idylla EGFR prototype assay was assessed by cell line dilution studies. A total of 10 ng was directly placed inside an Idylla EGFR prototype assay cartridge. Idylla results were compared with fragment length (exon 19 del) and Taqman assays.ResultsThe Idylla EGFR prototype assay showed an LOD of 1% mutant allele and yielded valid results in 74/76 (97.3%) samples, detecting all the mutant cases (n=32) identified by standard techniques; in addition, Idylla detected two low abundance EGFR exon 19 deletions and two G719X exon 18 point mutations, not covered by our standard reference method.ConclusionsIdylla EGFR prototype assay is sensitive on extracted DNA and can reliably be applied on cytological samples, enabling implementation of EGFR testing even in less experienced diagnostic units.
- Published
- 2016
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