Progress and challenges of sequencing and analyzing circulating tumor cells

Abstract

Circulating tumor cells (CTCs) slough off primary tumor tissues and are swept away by the circulatory system. These CTCs can remain in circulation or colonize new sites, forming metastatic clones in distant organs. Recently, CTC analyses have been successfully used as effective clinical tools to monitor tumor progression and prognosis. With advances in next-generation sequencing (NGS) and single-cell sequencing (SCS) technologies, scientists can obtain the complete genome of a CTC and compare it with corresponding primary and metastatic tumors. CTC sequencing has been successfully applied to monitor genomic variations in metastatic and recurrent tumors, infer tumor evolution during treatment, and examine gene expression as well as the mechanism of the epithelial-mesenchymal transition. However, compared with cancer biopsy sequencing and circulating tumor DNA sequencing, the sequencing of CTC genomes and transcriptomes is more complex and technically difficult. Challenges include enriching pure tumor cells from a background of white blood cells, isolating and collecting cells without damaging or losing DNA and RNA, obtaining unbiased and even whole-genome and transcriptome amplification material, and accurately analyzing CTC sequencing data. Here, we review and summarize recent studies using NGS on CTCs. We mainly focus on CTC genome and transcriptome sequencing and the biological and potential clinical applications of these methodologies. Finally, we discuss challenges and future perspectives of CTC sequencing.

Keywords: Circulating tumor cell; Next-generation sequencing; Single-cell sequencing.

Fig. 1

The workflow of circulating tumor cell sequencing

Fig. 2

Biological and potential clinical applications of CTC sequencing. a The CNV pattern of CTCs in a prostate cancer patient changed under therapeutic pressure. b CTC sequencing served as an efficient tool to uncover biological insights concerning tumor evolution. c RNA sequencing of CTCs enabled the exploration of metastasis-related pathways

Fig. 3

CTC sequencing biases introduced during amplification and library preparation. a Allele drop out (ADO) during genome amplification leads to the loss of the detection of somatic mutant alleles in CTCs, and false-positive results can also be introduced into both WGA and library processes. b Chimeras will lead to artificial CNV and SV detection in CTC sequencing. c The limitations of the WGA method may lead to low genome coverage or uneven read distribution effects when compared to bulk tumor sequencing