Performance of an RNA-Based Next-Generation Sequencing Assay for Combined Detection of Clinically Actionable Fusions and Hotspot Mutations in NSCLC

Introduction: With its expanding list of approved and emerging therapeutic indications, NSCLC is the exemplar tumor type requiring upfront assessment of several biomarkers to guide clinical management. Next-generation sequencing allows identification of different types of molecular alterations, each with specific analytical challenges. Library preparation using parallel DNA and RNA workflows can overcome most of them, but it increases complexity of laboratory operations, turnaround time, and costs. We describe the performance characteristics of a 15-gene RNA panel on the basis of anchored multiplex polymerase chain reaction for combined detection of clinically relevant oncogenic fusion transcripts and hotspot small variants... Conclusions: This ultrafocused RNA–next-generation sequencing assay offers an advantageous option with single unified workflow for simultaneous detection of clinically relevant hotspot mutations and fusions in NSCLC, focusing on actionable gene targets. READ ARTICLE

JTO Clinical and Research Reports DOI:10.1016/j.jtocrr.2022.100276

Authors: Patrice Desmeules, Dominique K. Boudreau, Nathalie Bastien, Marie-Chloé Boulanger, Yohan Bossé, Philippe Joubert, Christian Couture

A systematic review and meta-analysis of the adequacy of endobronchial ultrasound transbronchial needle aspiration for next-generation sequencing in patients with non-small cell lung cancer

Objectives: Next-generation sequencing (NGS) is able to identify targetable mutations to guide therapy and endobronchial ultrasound transbronchial needle aspiration (EBUS-TBNA) offers a potential route to routinely obtain specimens for analysis. However, the suitability of EBUS-TBNA samples for NGS remains uncertain... Conclusion: EBUS-TBNA was associated with a high yield for NGS and the success of EBUS-TBNA sampling for NGS was proportional to the number of passes. READ ARTICLE

Lung Cancer DOI:10.1016/j.lungcan.2022.01.018

Authors: Joseph J Zhao, Hiang Ping Chan, Yu Yang Soon, Yiqing Huang, Ross A Soo, Adrian C L Kee

The role of next-generation sequencing in detecting gene FUSIONS with KNOWN and UNKNOWN partners: A single-center experience with methodologies’ integration

Aims: Next-generation sequencing (NGS) is becoming a new gold standard for determining molecular predictive biomarkers. This study aimed to evaluate the reliability of NGS in detecting gene fusions, focusing on comparing gene fusions with known and unknown partners... Conclusions: NGS is a reliable method for detecting gene fusions with known partners, but it is less accurate in identifying gene fusions with unknown partners, for which further analyses (such as FISH) are required. READ ARTICLE

Human Pathology DOI:10.1016/j.humpath.2022.02.005

Authors: Andrea Ambrosini-Spaltro, Anna Farnedi, Daniele Calistri, Claudia Rengucci, Giovanna Prisinzano, Elisa Chiadini, Laura Capelli, Davide Angeli, Chiara Bennati, Mirca Valli, Giovanni De Luca, Dora Caruso, Paola Ulivi, Giulio Rossi

Review PaperKirk SmithFebruary 15, 2022Next-generation sequencing, FISH, IHC, gene fusion, partner, NSCLC

Clinical Outcomes for Plasma-Based Comprehensive Genomic Profiling Versus Standard-of-Care Tissue Testing in Advanced Non–Small Cell Lung Cancer

Background: Somatic genomic testing is recommended by numerous expert guidelines to inform targeted therapy treatment for patients with advanced nonsquamous non–small cell lung cancer (aNSCLC). The NILE study was a prospective observational study that demonstrated noninferiority of cell-free circulating tumor DNA (cfDNA)-based tumor genotyping compared to tissue-based genotyping to find targetable genomic alterations in patients with newly diagnosed nonsquamous aNSCLC. As the cohort has matured, clinical outcomes data can now be analyzed... Conclusions: cfDNA detects guideline-recommended biomarkers at a rate similar to tissue genotyping, and therapeutic outcomes based on plasma-based comprehensive genomic profiling are comparable to published targeted therapy outcomes with tissue profiling, even in community-based centers. READ ARTICLE

Clinical Lung Cancer DOI:10.1016/j.cllc.2021.10.001

Authors: Ray D. Page, Leylah M. Drusbosky, Hiba Dada, Victoria M. Raymond, Davey B. Daniel, Stephen G. Divers, Karen L. Reckamp, Miguel A. Villalona-Calero, Daniel Dix, Justin I. Odegaard, Richard B. Lanman, Vassiliki, A. Papadimitrakopoulou, Natasha B. Leighl

Blood First Assay Screening Trial (BFAST) in Treatment-Naive Advanced or Metastatic NSCLC: Initial Results of the Phase 2 ALK-Positive Cohort

Introduction: The Blood First Assay Screening Trial is an ongoing open-label, multicohort study, prospectively evaluating the relationship between blood-based next-generation sequencing (NGS) detection of actionable genetic alterations and activity of targeted therapies or immunotherapy in treatment-naive advanced or metastatic NSCLC. We present data from the ALK-positive cohort. Conclusions: These results reveal the clinical application of blood-based NGS as a method to inform clinical decision-making in ALK-positive NSCLC. READ ARTICLE

Journal of Thoracic Oncology DOI:10.1016/j.jtho.2021.07.008

Authors: Rafal Dziadziuszko, Tony Mok, Solange Peters, Ji-Youn Han, Jorge Alatorre-Alexander,
Natasha Leighl, Virote Sriuranpong, Maurice Pérol, Gilberto de Castro Junior, Ernest Nadal, Filippo de Marinis, Osvaldo Arén Frontera, Daniel S.W. Tan, Dae Ho Lee, Hye Ryun Kim, Mark Yan, Todd Riehl, Erica Schleifman, Sarah M. Paul, Simonetta Mocci, Rajesh Patel, Zoe June Assaf, David S. Shames, Michael S. Mathisen, Shirish M. Gadgeel

Clinical Relevance of an Amplicon-Based Liquid Biopsy for Detecting ALK and ROS1 Fusion and Resistance Mutations in Patients With Non–Small-Cell Lung Cancer

PURPOSE Liquid biopsy specimen genomic profiling is integrated in non–small-cell lung cancer (NSCLC) guidelines; however, data on the clinical relevance for ALK/ROS1 alterations are scarce. We evaluated the clinical utility of a targeted amplicon-based assay in a large prospective cohort of patients with ALK/ROS1-positive NSCLC and its impact on outcomes. CONCLUSION Next-generation, targeted, amplicon-based sequencing for liquid biopsy specimen profiling provides clinically relevant detection of ALK/ROS1 fusions in TKI-naïve patients and allows for the identification of resistance mutations in patients treated with TKIs. Liquid biopsy specimens from patients treated with TKIs may affect clinical outcomes and capture heterogeneity of TKI resistance, supporting their role in selecting sequential therapy. READ ARTICLE

JCO Precision Oncology DOI:10.1200/PO.19.00281

Authors: Laura Mezquita, Aurélie Swalduz, Cécile Jovelet, Sandra Ortiz-Cuaran, Karen Howarth, David Planchard, Virginie Avrillon, Gonzalo Recondo, Solène Marteau, Jose Carlos Benitez, Frank De Kievit, Vincent Plagnol, Ludovic Lacroix, Luc Odier, Etienne Rouleau, Pierre Fournel, Caroline Caramella, Claire Tissot, Julien Adam, Samuel Woodhouse, Claudio Nicotra, Edouard Auclin, Jordi Remon, Clive Morris, Emma Green, Christophe Massard, Maurice Pérol, Luc Friboulet, Benjamin Besse, and Pierre Saintigny

Real-World OutcomesKirk SmithApril 2, 2021Next-generation sequencing, liquid biopsy, ALK, ROS1, resistance

Intergenic Breakpoints Identified by DNA Sequencing Confound Targetable Kinase Fusion Detection in NSCLC

Introduction: Next-generation sequencing (NGS) based on genomic DNA has been widely applied for gene rearrangement detection in patients with NSCLC. However, intergenic-breakpoint fusions, in which one or both genomic breakpoints localize to intergenic regions, confound kinase fusion detection. We evaluated the function of intergenic-breakpoint fusions with multiplex molecular testing approaches... Conclusions: Intergenic-breakpoint fusions detected by DNA sequencing confound kinase fusion detection in NSCLC, as functional fusion transcripts may be generated or not. Additional validation testing using RNA/protein assay should be performed in intergenic-breakpoint fusion cases to guide optimal treatment. READ ARTICLE

Journal of Thoracic Oncology DOI:10.1016/j.jtho.2020.02.023

Authors: Weihua Li, Yutao Liu, Wenbin Li, Li Chen, Jianming Ying

Optimizing Mutation and Fusion Detection in NSCLC by Sequential DNA and RNA Sequencing

Introduction: Frequently, patients with locally advanced or metastatic NSCLC are screened for mutations and fusions. In most laboratories, molecular workup includes a multitude of tests: immunohistochemistry (ALK, ROS1, and programmed death-ligand 1 testing), DNA sequencing, in situ hybridization for fusion, and amplification detection. With the fast-emerging new drugs targeting specific fusions and exon-skipping events, this procedure harbors a growing risk of tissue exhaustion... Conclusions: We conclude that sequentially combining DNA NGS and RNA NGS is the most efficient strategy for mutation and fusion detection in smoking-associated NSCLC, whereas for never smokers we recommend a parallel approach. This approach was shown to be feasible on small tissue samples including for cytology tests, can drastically reduce the complexity and cost of molecular workup, and also provides flexibility in the constantly evolving landscape of actionable targets in NSCLC. READ ARTICLE

Journal of Thoracic Oncology DOI:10.1016/j.jtho.2020.01.019

Authors: Danielle Cohen, Liesbeth M. Hondelink, Nienke Solleveld-Westerink, Sandra M. Uljee, Dina Ruano, Anne-Marie Cleton-Jansen, Jan H. von der Thüsen, S. Rajen S. Ramai, Pieter E. Postmus, Jacob F. Graadt van Roggen, Bart P. C. Hoppe, Pieter C. Clahsen, Klaartje W. Maas, Els J. M. Ahsmann, Alexandraten Heuvel, Frank Smedts, Ronald N. van Rossem,Tomvan Wezel

Chr2 30297612-ALK, A Novel Intergenic Fusion With Exon18 of ALK, Responds to Crizotinib

Anaplastic lymphoma kinase (ALK) fusion is an important molecular subtype of non–small-cell lung cancer, and patients who have ALK fusion can be highly sensitive to ALK tyrosine kinase inhibitors. Different ALK fusions showed different sensitivities to ALK tyrosine kinase inhibitors. EML4-ALK (E6; A18) fusion showed resistance to crizotinib in a 48-year-old male patient with lung adenocarcinoma. READ ARTICLE

Clinical Lung Cancer DOI:10.1016/j.cllc.2020.04.014

Authors: Xiaofeng Chen, Guohong Zhao, Peilin Zhong, Min Zhang, Rongrong Chen, Derong Zhang

Biomarkers in Lung Cancer

Key Points: Biomarkers can be used for risk assessment, detection, diagnosis, and prognosis and to personalize treatment in lung cancer. Clinically useful biomarkers for selection of high-risk patients for lung cancer screening and to differentiate early lung cancer from benign pulmonary nodules are needed. Biomarkers for nodule management and determination of high-risk groups for lung cancer screening are at all phases of development, from discovery to clinical utility studies. Current trends in lung cancer biomarker development include the integration of clinical and radiologic features with molecular biomarkers, the application of artificial intelligence to molecular and imaging biomarker development, the use highly sensitive technologies such as next-generation sequencing for molecular exploration, and a commitment to high-quality clinical validation and utility studies. READ ARTICLE

Clinics in Chest Medicine. DOI: 10.1016/j.ccm.2019.10.004

Authors: Catherine R. Sears, Peter J. Mazzone

P109-32 Concurrent Genomic Alterations in ALK-Rearranged Non-Small Cell Lung Cancer Patients

Background: Recent progress in genomic analysis using next-generation sequencing (NGS) has enabled the comprehensive detection of targetable alterations in non-small cell lung cancer (NSCLC) patients. As the detection of ALK gene fusions is being established by NGS, identification of concurrent alterations will lead to better characterization of the molecular landscape of ALK-rearranged patients. Conclusion: We have studied the presence of ALK fusion genes with a novel NGS panel that showed excellent correlation with standard techniques. ALK fusions can be interpreted as early strong drivers to carcinogenesis due to the low frequency of concurrent alterations. It remains to determine the clinical impact of these alterations in larger series. READ ARTICLE

Journal of Thoracic Oncology DOI:10.1016/j.jtho.2019.08.1061

Authors: S. Clavé, M. Salido, J. Gibert, M. Hardy-Werbin, E. Weingartner, J. Hernandez, D. Nichol, P. Rocha, X. Riera, R. Blanco, J. Bosch-Barrera, Á. Taus, L. Pijuan, B. Bellosillo, E. Arriola

Next-generation Sequencing Identified a Novel WDPCP-ALK Fusion Sensitive to Crizotinib in Lung Adenocarcinoma

Anaplastic lymphoma kinase (ALK) fusion has been identified in 3% to 7% of patients with non–small-cell lung cancer (NSCLC), and such patients benefit greatly from ALK tyrosine kinase inhibitors (ALK-TKIs). The most common ALK fusion is echinoderm microtubule-associated protein-like 4 (EML4)-ALK, and several variants that are generally sensitive to crizotinib exist.1 In recent years, other fusion types sensitive to crizotinib, such as DYSF&ITGAV-ALK, BCL11A-ALK, and BIRC6-ALK, have been discovered by next-generation sequencing (NGS).2, 3, 4, 5, 6 However, a WD (Trp-Asp) planar cell polarity (PCP) effector gene (WDPCP)-ALK fusion has not been previously published. Here, we report this novel WDPCP-ALK fusion, which is sensitive to crizotinib, in a patient with lung adenocarcinoma. READ ARTICLE

Clinical Lung Cancer DOI:10.1016/j.cllc.2019.06.001

Authors: Zhen He, Xuan Wu, Shuxiang Ma, Cuicui Zhang, Zhe Zhang, Shuai Wang, Sheng Yu, Qiming Wang

Case StudyKirk SmithSeptember 1, 2019C1156Y, Crizotinib, Next-generation sequencing, Sensitivity, WDPCP-ALK