Table 3.

Next-generation sequencing of circulating cell-free DNA: clinical utility.

Patients (Samples)	Technology	Findings and Potential Clinical Implications	Ref.
Static cf/ctDNA next-generation sequencing analysis
21,807 (>50 advanced cancer types)	tNGS	Driver gene cfDNA mutation profiles were similar to tumor NGS, while differences were attributed to clonal evolution over therapy leading to resistance	[70]
1422 (sub-study, 21 tumor types)	tNGS, WGS, WGBS	Sensitivity for 12 cancers including CRC was 76% and 74% for stage I-III CRC	NCT02889978 [71]
1397 (advanced CRC)	tNGS	Mutation frequencies in ctDNA were similar to tissue, and multiple distinct resistant mutations were identified in single patients	[19]
1005 (8 cancer types)	CancerSEEK	Sensitivity was 65% and stage-dependent for CRC, suggesting the need for improvement before clinical applicability	[72]
100 (TARGET study, diverse advanced cancers, 23 CRC)	tNGS	Druggable mutations were identified in 41/100 pts, 11/41 received matched therapy and all 11 achieved PR or stable disease	[22]
80 pts	WGS	Recurrent CNVs were identified in multiple chromosomal regions and correlated with stage and prognosis	[73]
Consecutive liquid biopsies before and after systemic therapy
261 (ASPECCT study, plasma samples before and after panitumumab)	tNGS	Baseline high RAS mutant allele frequency and EGFR pathway mutations were adverse prognostic factors, while tumor mutational burden increased over time This study suggests potential utility for primary and secondary decision-making	[74]
238 (ASPECCT study, plasma samples before and after panitumumab)	tNGS	79% of baseline samples were WT and 21% mutant RAS (associated with worse outcomes), while 32% of baseline-WT tumors had emergent RAS mutations	[75]
53 (159 serial samples over chemotherapy)	tNGS	Mutational concordance between tumor and cfDNA was 92.3%, while cfDNA levels were predictive of clinical response	[76]
39 various metastatic cancers, 12 CRC (159 total serial samples over targeted therapy)	tNGS	Monitoring of plasma mutation allele identified potential clonal responses to targeted therapy associated with progression, suggesting potential prognostic and predictive utility	[77]

Abbreviations: cell-free DNA (cfDNA), circukating tumor DNA (ctDNA), colorectal cacner (CRC), next-generation sequencing (NGS), partial response (PR), targeted NGS (tNGS), whole-genome bisulfite sequencing (WGBS), whole-genome sequencing (WGS), wild-type (WT).