CUT-PCR: CRISPR-mediated, ultrasensitive detection of target DNA using PCR

Area of Science:

• Biotechnology
• Molecular Biology
• Genomics

Background:

• Circulating tumor DNA (ctDNA) is a promising biomarker for early cancer detection.
• Current ctDNA enrichment methods lack sufficient sensitivity for early-stage diagnosis.
• Detecting minute amounts of tumor DNA amidst wild-type DNA remains a challenge.

Purpose of the Study:

• To develop a novel, ultrasensitive method for detecting ctDNA.
• To improve the sensitivity and accuracy of cancer diagnosis using ctDNA.
• To establish a broadly applicable technique for early cancer detection.

Main Methods:

• Development of CRISPR-mediated, Ultrasensitive detection of Target DNA (CUT-PCR) technology.
• Utilizing CRISPR endonucleases (e.g., SpCas9, FnCpf1) to eliminate wild-type DNA sequences.
• Integration of CUT-PCR with targeted deep sequencing for enhanced analysis.

Main Results:

• CUT-PCR demonstrates applicability to 80% of cancer-linked mutations in the COSMIC database.
• Achieved high sensitivity (<0.01%) and accuracy in detecting oncogenes.
• Outperformed conventional targeted deep sequencing in sensitivity and accuracy.
• Successfully detected oncogenic mutations in ctDNA from colorectal cancer patients.

Conclusions:

• CUT-PCR offers a significant advancement in ultrasensitive ctDNA detection.
• The method shows potential for early diagnosis of various cancers.
• This technique could revolutionize non-invasive cancer diagnostics.