A DNA alteration and methylation co-detection method for clinical purpose

Area of Science:

• Molecular Biology
• Genomics
• Epigenetics

Background:

• Current genomic and methylation analyses require separate assays, complicating workflows and limiting low-input sample use.
• Cell-free DNA (cfDNA) analysis is challenged by low sample input and the need for multi-omic data.

Purpose of the Study:

• To develop a streamlined assay for simultaneous mutation and methylation profiling.
• To evaluate the performance of this integrated assay in various clinical contexts, including cfDNA analysis.

Main Methods:

• Developed a novel method using mutation-protective strand synthesis with modified deoxycytidine triphosphates.
• Validated the approach using whole-genome sequencing of cell lines and targeted sequencing of clinical samples.
• Implemented the assay on a quantitative PCR (qPCR) platform.

Main Results:

• Demonstrated high concordance with standard DNA sequencing and methylation assays.
• Achieved a modest ~12% improvement in circulating tumor DNA (ctDNA) detection in lung cancer patients.
• Revealed links between homologous recombination repair (HRR) gene function and homologous recombination deficiency (HRD) in gynecologic cancers.
• Showcased high performance on qPCR with a 0.5% limit of detection.

Conclusions:

• The integrated mutation and methylation profiling approach simplifies clinical workflows and enhances cfDNA analysis.
• This method shows promise for improved ctDNA detection and understanding cancer biology.
• The adaptability and performance of this assay suggest significant potential for advancing clinical diagnostics.