Toma Os-Seq: A Clinically Oriented Targeted Resequencing Method For Detecting All Mutation Classes In Dna Isolated From Ffpe, Fnas And Plasma

Critical genetic errors, namely specific mutations, are required for the development and maintenance of cancer. Molecular diagnostics that leverage the breadth of next-generation sequencing (NGS) to identify this mutational landscape are therefore becoming an important clinical tool in the precision treatment of cancer. However, the widespread clinical adoption of NGS technologies is hampered not only by the challenges of assaying from various clinical matrices, but also by the complexity of the workflows utilized to create, analyze, and annotate an NGS library. To address these challenges, an adaptation of oligo-selective sequencing (OS-Seq) was developed with a workflow designed specifically for the requirements of the clinical laboratory. Termed TOMA OS-Seq)a panel of 96 genes - each with diagnostic or prognostic value - can be sequenced end-to-end from as little as 1 ng of input material, enabling targeted library construction from diverse sources such as fine-needle aspirates (FNAs), plasma as well as formalin-fixed paraffin embedded (FFPE) tissue. The minimized number of high-efficiency steps reduces the requirement for PCR, allowing the identification of copy number alterations (CNAs), re-arrangements, insertions and deletions (indels), as well as single nucleotide variants (SNVs) from these matrices. The performance of TOMA OS-Seq) was assessed using a panel of cell lines harboring known variants in all classes of mutations. Specifically, comparison with a 12-gene droplet digital PCR (ddPCR) based test (the Amplinome Test) demonstrated a high correlation (R2 u003e 0.95) of CNA measurements via TOMA OS-Seq) with ddPCR, when further challenged with FNAs. The application of TOMA OS-Seq) in assaying all mutation classes in circulating tumor DNA from plasma demonstrates the robustness and flexibility of this platform as clinically oriented targeted resequencing method. Citation Format: Yosr Bouhlal, Alexander McKenzie, Austin P. So. TOMA OS-Seq: A clinically oriented targeted resequencing method for detecting all mutation classes in DNA isolated from FFPE, FNAs and plasma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4885. doi:10.1158/1538-7445.AM2015-4885