Abstract 5704: Ultrasensitive mutational analysis in cell-freeDNA by targeted next-generation sequencing with molecular tags

Abstract

Circulating cell-free DNA (cfDNA) in plasma offers a non-invasive approach to monitor tumor molecular profiling in real-time at multiple time-points, detection of emerging genomic alterations associated with drug resistance and clarifying cancer prognosis and diagnosis of cancer recurrence or progression. Therefore, it is crucial to improve our current NGS technology to detect low frequency alleles in cfDNA.

There are many technical challenges in NGS technology, including base errors generated from library preparation and sequencing, making it hard to distinguish real mutations from false positives produced from amplification sequencing and base-calling processes.

We have adopted a more sensitive approach to overcome this challenge. We used short, random DNA sequences called unique molecular tags (UMTs) during library preparation to label input DNA. This technique can remove background errors, false positives during data analysis, providing high confident mutation calls with better sensitivity down to 0.1% (depends on DNA input amount).

We will present data showing validation of this NGS platform using commercially available DNA control, archival FFPE tissue and cfDNA matched samples and demonstrate robust sensitivity and specificity by using the off-the-shelf Archer Reveal ctDNA panel for Illumina covers 28 oncogenes. This molecular-tagged NGS panel can detect mutations down to 0.1% allelic frequency. This robust NGS platform is being used in our lab to analyze cfDNA samples, enabling the development of a non-invasive method to overcome existing challenges to provide molecular understanding of patient’s tumor evolution in real time, and aid in the development of personalized therapies for cancer patients.

Citation Format: Nga Wan Rachel Tam, Irmina Diala, Xiaoji Chen, Walter Darbonne. Ultrasensitive mutational analysis in cell-freeDNA by targeted next-generation sequencing with molecular tags [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5704. doi:10.1158/1538-7445.AM2017-5704