Cell-free DNA (cfDNA) in the circulating blood plasma of patients with cancer contains tumour-derived DNA sequences that can serve as biomarkers for guiding therapy, for the monitoring of drug resistance, and for the early detection of cancers. However, the analysis of cfDNA for clinical diagnostic applications remains challenging because of the low concentrations of cfDNA, and because cfDNA is fragmented into short lengths and is susceptible to chemical damage. Barcodes of unique molecular identifiers have been implemented to overcome the intrinsic errors of next-generation sequencing, which is the prevailing method for highly multiplexed cfDNA analysis. However, a number of methodological and pre-analytical factors limit the clinical sensitivity of the cfDNA-based detection of cancers from liquid biopsies. In this Review, we describe the state-of-the-art technologies for cfDNA analysis, with emphasis on multiplexing strategies, and discuss outstanding biological and technical challenges that, if addressed, would substantially improve cancer diagnostics and patient care.
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We thank S. Skates for useful discussions regarding diagnostic economics and outcomes. L.N.K. was supported by NIH grant P01CA163222. A.A.P. was supported by NIH grants R01CA197486 and R01CA233364. D.Y.Z. was supported by NIH grants R01CA203964 and R01CA233364, and by CPRIT grant RP180147.
P.S., L.R.W. and A.A.P. are consultants for NuProbe USA. A.A.P. is a consultant for Binary Genomics and has equity ownership in the company. D.Y.Z. is a co-founder of, and holds significant equity in, NuProbe Global, Torus Biosystems and Pana Bio.
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Song, P., Wu, L.R., Yan, Y.H. et al. Limitations and opportunities of technologies for the analysis of cell-free DNA in cancer diagnostics. Nat. Biomed. Eng 6, 232–245 (2022). https://doi.org/10.1038/s41551-021-00837-3
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