Development of an integrable RNA based Gene Panel for Next-Generation Sequencing of Clinically Relevant Gene Fusions in Non-Small Cell Lung Cancer Patients

Non-small cell lung cancer (NSCLC) still represents the main cause of cancer mortality worldwide. However, targeted treatments have significantly improved the clinical outcome of those patients with non-squamous NSCLC harboring not only point mutations and indertions/deletions (indels), but also gene rearrangements. Currently, immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) are considered the "gold standard" methodologies for the identification of these genomic alterations but feature some technical and interpretative limitations. Therefore, the aim of this study is to validate a novel narrow custom next generation sequencing gene panel, termed SiRe gene fusion panel, able to detect gene rearrangements and other aberrations (such MET exon 14 skipping) for the administration of specific tyrosine kinase inhibitors

Diagnostic yield of rapid on-site evaluation transbronchial needle aspiration versus conventional transbronchial needle aspiration: a single center experience

Trans-bronchial needle aspiration allows lymph node sampling in several thoracic conditions; the ability of Rapid On-Site Evaluation (ROSE) to predict the final diagnosis in this setting has not been well characterized. We performed a retrospective study to establish the utility of ROSE in the diagnosis of thoracic diseases with mediastinal lymph node involvement. We retrospectively reviewed 297 patients with hilar-mediastinal lymph node enlargement detected at CT scan from January 2013 to April 2016. 201 patients underwent conventional TBNA; in 96 patients, TBNA procedure was performed by on-site presence of a team of pathologists and research morphologists. Lung neoplasms, sarcoidosis, infections and lymphoma were the most common diseases diagnosed with TBNA samples. TBNA simultaneously performed in combination with ROSE produced an increase in percentage of appropriate samples compared to single cTBNA (adequate samples cTBNA vs ROSE-TBNA: 73% vs 81%; p<0.05). Our observations indicate an increase in adequacy of fine needle aspirations and increased diagnostic yield in the ROSE group. In conclusions, ROSE may serve to reduce procedure time and enhance sample triaging therefore limiting the need for further invasive diagnostic testing

Microsatellite instability evaluation by automated microfluidic electrophoresis: an update

A significant proportion (∼15%) of colorectal cancer (CRC), either sporadic or arising in the setting of the hereditary non-polyposis colorectal carcinoma syndrome, features microsatellite instability (MSI). Five MSI loci, either mononucleotide or dinucleotide repeats (Bat25, Bat26, D2S123, D5S346 and D17S250), are included in the Bethesda panel and capillary electrophoresis represents the usual gold standard technique

Next generation sequencing techniques in liquid biopsy: focus on non-small cell lung cancer patients

The advent of genomic based personalized medicine has led to multiple advances in the molecular characterization of many tumor types, such as non-small cell lung cancer (NSCLC). NSCLC is diagnosed in most cases on small tissue samples that may be not always sufficient for EGFR mutational assessment to select patients for first and second generations' tyrosine kinase inhibitors (TKIs) therapy. In patients without tissue availability at presentation, the analysis of cell free DNA (cfDNA) derived from liquid biopsy samples, in particular from plasma, represent an established alternative to provide EGFR mutational testing for treatment decision making. In addition, a new paradigm for TKIs resistance management was recently approved by Food and Drug Administration, supporting the liquid biopsy based genotyping prior to tissue based genotyping for the detection of T790M mutation to select patients for third generation TKIs. In these settings, real time PCR (RT-PCR) and digital PCR 'targeted' methods, which detect known mutations by specific probes, have extensively been adopted. Taking into account the restricted reference range and the limited multiplexing power of these targeted methods, the performance of liquid biopsy analyses may be further improved by next generation sequencing (NGS). While most tissue based NGS genotyping is well established, liquid biopsy NGS application is challenging, requiring a careful validation of the whole process, from blood collection to variant calling. Here we review this evolving field, highlighting those methodological points that are crucial to accurately select NSCLC patients for TKIs treatment administration by NGS on cfDNA

Performance analysis of SiRe next-generation sequencing panel in diagnostic setting: focus on NSCLC routine samples

Following 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