A novel approach for next‐generation sequencing of circulating tumor cells. Issue 4 (28th February 2016)

• Record Type: Journal Article
• Authors: Yee, Stephanie S.; Lieberman, David B.; Blanchard, Tatiana; Rader, JulieAnn; Zhao, Jianhua; Troxel, Andrea B.; DeSloover, Daniel; Fox, Alan J.; Daber, Robert D.; Kakrecha, Bijal; Sukhadia, Shrey; Belka, George K.; DeMichele, Angela M.; Chodosh, Lewis A.; Morrissette, Jennifer J. D.; Carpenter, Erica L.
• Is Part Of: Molecular genetics & genomic medicine. Volume 4:Issue 4(2016)
• Page Start: 395
• Page End: 406
• Publication Date: 2016-02-28
• Subjects: Breast cancer -- circulating tumor cell -- liquid biopsy -- next‐generation sequencing -- personalized medicine -- whole genome amplification; Medical genetics -- Periodicals; Genomics -- Periodicals; 616.042
• DOI: 10.1002/mgg3.210
• Languages: English
• ISSNs: 2324-9269

Abstract

Abstract: Background: Next‐generation sequencing (NGS) of surgically resected solid tumor samples has become integral to personalized medicine approaches for cancer treatment and monitoring. Liquid biopsies, or the enrichment and characterization of circulating tumor cells (CTCs) from blood, can provide noninvasive detection of evolving tumor mutations to improve cancer patient care. However, the application of solid tumor NGS approaches to circulating tumor samples has been hampered by the low‐input DNA available from rare CTCs. Moreover, whole genome amplification (WGA) approaches used to generate sufficient input DNA are often incompatible with blood collection tube preservatives used to facilitate clinical sample batching. Methods: To address this, we have developed a novel approach combining tumor cell isolation from preserved blood with Repli‐G WGA and Illumina TruSeq Amplicon Cancer Panel‐based NGS. We purified cell pools ranging from 10 to 1000 cells from three different cell lines, and quantitatively demonstrate comparable quality of DNA extracted from preserved versus unpreserved samples. Results: Preservation and WGA were compatible with the generation of high‐quality libraries. Known point mutations and gene amplification were detected for libraries that had been prepared from amplified DNA from preserved blood. Conclusion: These spiking experiments provide proof of concept of a clinically applicable workflow for real‐time monitoring of patient tumor using noninvasive liquid biopsies. Abstract : The application of solid tumor next‐generation sequencing (NGS) approaches to circulating tumor samples has been hampered by the low‐input DNA available from rare circulating tumor cells (CTCs), and whole genome amplification (WGA) approaches used to generate sufficient input DNA are often incompatible with blood collection tube preservatives used to facilitate clinical sample batching. We developed a novel approach combining tumor cell isolation from preserved blood with Repli‐G WGA and Illumina TruSeq Amplicon Cancer Panel‐based NGS. We applied this approach to purified cell pools ranging from 10 to 1000 cells from three different cell lines to provide proof of concept of a clinically applicable workflow for real‐time monitoring of patient tumor using noninvasive liquid biopsies.