Single-Molecule Sequencing Enables Long Cell-Free DNA Detection and Direct Methylation Analysis for Cancer Patients.
Clin Chem
; 68(9): 1151-1163, 2022 09 01.
Article
em En
| MEDLINE
| ID: mdl-35587130
ABSTRACT
BACKGROUND:
Analysis of circulating tumor DNA has become increasingly important as a tool for cancer care. However, the focus of previous studies has been on short fragments of DNA. Also, bisulfite sequencing, a conventional approach for methylation analysis, causes DNA degradation, which is not ideal for the assessment of long DNA properties and methylation patterns. This study attempted to overcome such obstacles by single-molecule sequencing.METHODS:
Single-molecule real-time (SMRT) sequencing was used to sequence plasma DNA. We performed fragment size and direct methylation analysis for each molecule. A methylation score concerning single-molecule methylation patterns was used for cancer detection.RESULTS:
A substantial proportion of plasma DNA was longer than 1â kb with a median of 16% in hepatocellular carcinoma (HCC) patients, hepatitis B virus carriers, and healthy individuals. The longest plasma DNA molecule in the HCC patients was 39.8â kb. Tumoral cell-free DNA (cfDNA) was generally shorter than nontumoral cfDNA. The longest tumoral cfDNA was 13.6â kb. Tumoral cfDNA had lower methylation levels compared with nontumoral cfDNA (median 59.3% vs 76.9%). We developed and analyzed a metric reflecting single-molecule methylation patterns associated with cancer, named the HCC methylation score. HCC patients displayed significantly higher HCC methylation scores than those without HCC. Interestingly, compared to using short cfDNA (area under the receiver operating characteristic [ROC] curve, AUC 0.75), the use of long cfDNA molecules greatly enhanced the discriminatory power (AUC 0.91).CONCLUSIONS:
A previously unidentified long cfDNA population was revealed in cancer patients. The presence and direct methylation analysis of these molecules open new possibilities for cancer liquid biopsy.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Carcinoma Hepatocelular
/
Ácidos Nucleicos Livres
/
Neoplasias Hepáticas
Tipo de estudo:
Diagnostic_studies
/
Prognostic_studies
Limite:
Humans
Idioma:
En
Revista:
Clin Chem
Assunto da revista:
QUIMICA CLINICA
Ano de publicação:
2022
Tipo de documento:
Article
País de afiliação:
China