Abstract 215: Oncomineprecision assay - an integrated and automatic next generation sequencing workflow for both FFPE tissue and plasma based cancer research

Abstract

Introduction: Next generation sequencing technologies and platforms have been evolving rapidly over the last decade with improved throughput and cost-effectiveness. The application of various NGS assays using both genome wide or targeted sequencing has been widely employed in cancer research to characterize tumor biomarker profiles including SNV/indels, CNV, and fusion in various sample types. Here we will present the performance of OncomineTM Precision Assay developed at Thermo Fisher Scientific, which is compatible with both solid tumor and liquid biopsy sample types. It is the very first automated NGS assay developed with Ion Torrent Genexus system to provide a streamlined workflow directly from sample to results with minimal hands-on time. Methods: The assay employs a novel AmpliSeq™ HD library technology to provide optimal sensitivity and specificity for challenging sample types including degraded FFPE or cell-free nucleic acid from plasma. For library preparation, 10 ng DNA or RNA was used for FFPE and 20 ng cfTNA was used for plasma samples. Ion Torrent Genexus system provides an interface to guide the user to plan the run following a defined workflow according to sample type, and performs library preparation, templating and sequencing automatically. Results: The OncomineTM Precision Assay includes focused genetic content with clinical relevance including 45 hotpot genes (SNV/Indel), 14 CNVs, and 19 structural variants (fusion and exon variants). This study evaluated the assay performance using a set of controls for FFPE or cfDNA from positive cell line material or commercial sources such as Horizon discovery and SeraCare, which contain various SNV/Indels on several important gene loci (EGFR, KRAS, NRAS, BRAR, and PIK3CA), CNV (EGFR, MET amplification and pTEN deletion), and fusion variants across ALK, ROS1, RET, and NTRKs. The results showed that all the expected SNV variants were consistently detected with sensitivity at ≥95% for FFPE with 5% AF and ≥85% for cfDNA samples down to 0.1% LOD. For fusion variants and CNV targets including gain and loss, both sensitivity and PPV at ≥90% could be achieved for FFPE and cfTNA samples. Furthermore, a cohort of positive FFPE tissue was also analyzed and detected expected variants that had been previously confirmed through an orthogonal NGS test. Lastly, a set of matched FFPE and plasma samples was also evaluated using the assay and demonstrated high concordance between FFPE and plasma from each subject. Conclusion: Overall the study demonstrates that the OncomineTM Precision Assay provides an automated and sensitive NGS solution for efficient and effective mutation assessment using both FFPE and liquid biopsy samples. Citation Format: Ru Cao, Jian Gu, Jeff Schageman, Kris Lea, Priyanka Kshatriya, Amir Marcovitz, Paul Williams, Bo Ding, Khalid Hanif, Kelli Bramlett. Oncomineprecision assay - an integrated and automatic next generation sequencing workflow for both FFPE tissue and plasma based cancer research [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 215.

Implementation of an ISO15189 accredited next-generation sequencing service with the fully automated Ion Torrent Genexus: the experience of a clinical diagnostic laboratory

“…The Genexus streamlines the workflow and can deliver sequencing results in a single day, while meeting national and international guidelines and standards. 17 The OR software was successfully integrated allowing NGS reporting without the need for bioinformatics expertise. Bespoke report templates were curated and aligned to local requirements, thus facilitating the linking of variants to labels guidelines and clinical trials.…”

“…The Genexus system was chosen as it did not require user expertise in bioinformatics and reporting; it facilitated a minimal sample input volume and flexible run sizes with a cost per assay model instead of cost per run. The Genexus streamlines the workflow and can deliver sequencing results in a single day, while meeting national and international guidelines and standards 17…”

“…16 In this study, we set out to introduce a novel, fully automated NGS workflow, with a sequencing TAT (24 hours) currently not attainable in this country. The Ion Torrent Genexus is recognised as the first 'turnkey' NGS solution that streamlines the workflow and can deliver sequencing results in a single day, 17 it was selected as the solution for this laboratory as it addresses the barriers listed above with an automated specimen to report workflow. The Genexus does not require a high level of user expertise in bioinformatics and reporting; it facilitates a minimal nucleic acid input volume of approximately 10 ng, it has a multiplexing capability of up to 32 library preparations in a single run while also allowing run sizes of 2-4 samples using a cost per assay model instead of cost per run.…”

Implementation of an ISO 15189 accredited next generation sequencing service for cell-free total nucleic acid (cfTNA) analysis to facilitate driver mutation reporting in blood: the experience of a clinical diagnostic laboratory

“…42 The Genexus may also be used for cfTNA isolated from plasma without requiring a high level of user expertise in bioinformatics and reporting. 43 It facilitates a minimal nucleic acid input volume of approximately 5 ng, with run sizes of 1-5 cfTNA samples, using a cost per assay instead of a cost per run model.…”