Detection of Point Mutation in Individual Living Circulating Malignant Cells by Targeting Molecular Beacon Delivery

Area of Science:

• Biomedical Engineering
• Molecular Biology
• Oncology

Background:

• Single-base substitutions are key drivers of cancer and drug resistance.
• Current single-cell mutation detection methods are costly and time-consuming.
• Accurate detection of tumor heterogeneity is crucial for personalized cancer therapy.

Purpose of the Study:

• To develop an efficient and specific method for detecting single-base mutations in circulating malignant cells (CMCs) at the single-cell level.
• To enable real-time profiling of tumor heterogeneity in cancer patients.
• To establish a non-invasive liquid biopsy strategy for in situ tumor genotyping.

Main Methods:

• Development of an aptamer-functionalized triple-targeting nanoprobe.
• Integration of AT11 and R13 aptamers with hyaluronic acid for targeted delivery.
• Utilizing molecular beacons for hybridization with target mRNAs (GAPDH, wild-type EGFR, L858R-mutated EGFR) in living CMCs.
• Comparison of fluorescence intensities to determine EGFR L858R mutation status.

Main Results:

• The nanoprobe achieved efficient and specific delivery of molecular beacons into CMCs in whole blood.
• Single-cell resolution profiling of EGFR L858R point mutation status was successfully demonstrated.
• Patient-specific tumor heterogeneity was mapped at the individual patient level.

Conclusions:

• The developed nanoprobe offers a versatile, non-invasive liquid biopsy strategy for in situ tumor genotyping.
• This approach allows for precise mapping of tumor heterogeneity at single-cell resolution.
• The technology has wide applicability for cancer monitoring and therapeutic guidance.