Abstract 4148: Sensitive detection of tumor nucleic acids in plasma by mutation-enriched next generation sequencing.

Abstract

Abstract We have demonstrated a novel somatic mutation enrichment methodology that allows DNA sequencing to work beyond its usual limit of detection to accurately profile solid tumors by detecting their mutation signature in plasma, even when the tumor DNA is present in abundances below 0.01%. Next Generation DNA Sequencing (NGS) is becoming the new standard for mutational profiling of tumor tissue, due to its flexibility, speed, and decreasing cost. While generally exceptional in performance, NGS suffers from a sequencing error rate of 0.1% - 1%, largely due to amplification induced artifacts in its workflow. While this does not constitute a significant problem in application of NGS to sequencing of most tumour tissues, it makes NGS impractical as a method to search for low abundance mutation signatures in plasma samples. Numerous publications have shown the presence of tumor signatures in the cell-free DNA (cfDNA) circulating in plasma, but concordance between the tumor signature and the plasma signature has been limited. This is likely due to limitations in the detection technologies used to search for cfDNA in plasma. To maximize concordance between plasma and tissue, it will be essential that sensitivities reaching 0.01% and below (as little as a single tumor mutant allele per sample) be achieved, and ideally that multiple mutational hot spots be analyzed to maximize the chance of detection. Current technologies are incapable of such sensitivity over a large number of mutation loci. We have developed a novel electrophoretic method that can enrich nucleic acid samples over 1,000,000-fold for up to 100 somatic mutations, enabling reliable profiling of samples containing as little as 0.01% mutant. By enriching nucleic acid samples for specific targets prior to amplification and sequencing, we enable the use of NGS in plasma-based mutation detection and profiling. We present data demonstrating highly sensitive multiplexed mutation detection in plasma samples, demonstrating 0.01% sensitivity. We also report on concordance studies between mutation incidence in tissue and matched plasma in human samples. Citation Format: Andre Marziali. Sensitive detection of tumor nucleic acids in plasma by mutation-enriched next generation sequencing. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4148. doi:10.1158/1538-7445.AM2013-4148