Cytopathology and Predictive Molecular Pathology
Coordinator
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Prof. Giancarlo Troncone Full Professor of Anatomic Pathology Head of the Department of Public Health |
Research lines
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The research activity of the "Cytopathology and Predictive Molecular Pathology" unit focuses on the following research lines: • Evaluation of the mutational status of clinical relevant predicitve biomarkers of response to targeted treatment in different solid tumors. The research activity is aimed to define the assess the mutational status of specific genes (KRAS, NRAS, EGFR, PIK3CA, BRAF, BRCA) able to predict responsiveness to targeted treatments, represented by monoclonal antibodies (mAB) and tyrosine kinase inhibitors (TKIs), in different solid tumors (metastatic colorectal cancer, lung cancer, ovarian cancer, breast cancer, melanoma, gastrointestinal stromal tumor). The activity is also focused on the optimization of novel molecular techniques, such as next generation sequencing (NGS) approach. • Evaluation of molecular alterations to refine the risk of malignancy of uncertain morphological thyroid fine needle aspiration diagnostic classes. As part of the TIRNET project, funded by the Campania region, the unit is oriented in the evaluation of molecular alterations useful for the diagnostic stratification of thyroid nodules classified as indeterminate (TIR3A / B). The unit, which performs the role of hub center in the molecular analysis of nodules with indeterminate morphology within the regional network, has activated a diagnostic-analytical protocol using technologies with a high diagnostic and clinical impact (RT-PCR and NGS). • Liquid Biopsy for the molecular profiling of cancer patients. In this context, the unit aims at the analytical validation of biological fluids (urine, blood, sputum, cerebrospinal fluid) as alternative sources of tumoral nucleic acids for the detection of clinically relevant gene alterations to select cancer patients for treatment with targeted therapies. In particular, the use of peripheral blood (liquid biopsy) for the molecular analysis of patients with advanced lung cancer. The research protocol is aimed to optimize the analytical management workflow of the sample through the implementation of validated and controlled procedures (ranging from nucleic acid extraction to molecular analysis conducted by using NGS platforms) aimed to maximize the adoption of cell free DNA (cfDNA) in clinical practice. Another important point is represented by the design, development and validation of diagnostic devices for NGS platforms for the analysis of clinically relevant mutations in cancer patients. The research activity is aimed at the definition and development of customized gene panels designed to increase the sensitivity and analytical specificity of the molecular analysis of predictive biomarkers of response to targeted treatments. The SiRe gene panel, able to detect 568 clinically relevant mutations within KRAS, NRAS, PIK3CA, EGFR, BRAF, KIT, PDGFRA genes, has been validated from a technical-analytical and clinical point of view to allow tissue and blood samples analysis. The device received RUO certification and was the subject of a patent before being used in clinical practice. In this context, a custom, RNA-based gene panel was also created, able to analyze on the NGS platform gene fusions in ALK, ROS1, RET, NTRK genes and the MET exon 14 skipping alterations that play a leading role in the clinical management of patients with non-small cell lung cancer (NSCLC).
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Funding deriving from competitive tenders.
If so, indicate which ones.
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POR Campania FESR 2014/2020 progetto "Sviluppo di Approcci Terapeutici Innovativi per Patologie Neoplastiche Resistenti ai Trattamenti – SATIN. |
Instrumental Platforms
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The Cytopathology and Predictive Molecular Pathology unit employs four different NGS platforms (Ion Torrent PGM ™, Ion Torrent S5, Ion Torrent S5 Plus; Genexus) and two automatic library preparers for NGS analysis (Ion Chef ™). In addition, the Unit has an automatic extractor (Qia-Simphony) used to standardize the cfDNA extraction phase of liquid biopsy samples. The Unit also adopts three thermal cyclers (MiniAmp) used in the working phases of tissue and plasma samples, a nanofluidic platform (Tape Station4200) for the quantitative and quantitative evaluation of the nucleic acids extracted from the samples prior to the NGS analysis; a massive and parallel hybridization platform with fluorescent probes (n-Counter) used in the evaluation of gene expression profiles. Finally, the Unit has two RT-qPCR platforms (Idylla ™ and EasyPGX) which allow the full automation of the real time analysis of nucleic acids. |
Third Mission |
Spin-off GENEDIN s.r.l. 1. From 2017 - in progress: Spin - Off of the Department of Public Health of the University of Naples Federico II GENEDIN s.r.l. with registered office in Via Giovanni Devoti 14 (Rome, Italy). The first diagnostic device born in this spin-off was represented by a panel of oligonucleotide probes for the characterization of genes that predict the response to next-generation drug treatments for patients with metastatic colorectal cancer, non-small cell lung cancer, melanoma and gastrointestinal stromal tumors ( certified for in vitro diagnostics by the European community - CE - IVD). Trademark Registration 1. The SiRe® brand that identifies products marketed by the Spin - Off GENEDIN s.r.l., developed by enhancing the intellectual products of the Predictive Molecular Pathology laboratory of the Department of Public Health is owned by 50% of the public proposing partners of the spin - off.
Development and validation of gene panels for the massive and parallel sequencing of nucleic acids extracted from fine needle aspiration cytology samples carried out in patients with thyroid nodules for the definition of the risk of malignancy. The project financed by the Campania Region with regional law No. 24 of 29/12/2005, led to the creation of the TIRNET regional network to carry out molecular tests in thyroid cytopathology. Through an IT platform (www.tirnet.it) some of the main hospitals in Campania (AORN Pascale, Cardarelli, Ospedale dei Colli, AOU Vanvitelli, ASL1-Ospedale del Mare, Moscati di Avellino and "dell 'Immacolata" Hospital of Sapri ) periodically send biological material from thyroid undetermined nodules to the Molecular Cytopathology laboratories of the Department of Public Health of the University of Naples Federico II to perform molecular tests (detection of mutations and fusions). In a first phase, an RT-PCR test was validated for the evaluation of the most frequent alterations in thyroid neoplasms (BRAF, RAS, RET / PTC, PAX8 / PPARg). Subsequently, a preliminary panel was designed and validated for the evaluation of a greater number of alterations (BRAF, EIF1AX, GNAS, HRAS, IDH1, KRAS, NF2, NRAS, PIK3CA, PPM1D, PTEN, RET, DICER1, CHEK2, TERT promoter ), using next-generation sequencing technologies. Thanks to the partnership with Thermofisher Scientific, the design and validation of an improved version of the aforementioned panel is underway, which will allow not only the evaluation of a greater number of gene alterations but also the evaluation of the mRNA expression expressed by thyroid cells both in physiological and pathological conditions. |
Personnel of the unit |
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