Rapid Multiplex Strip Test for the Detection of Circulating Tumor DNA Mutations for Liquid Biopsy Applications

In the era of personalized medicine, molecular profiling of patient tumors has become the standard practice, especially for patients with advanced disease. Activating point mutations of the KRAS proto-oncogene are clinically relevant for many types of cancer, including colorectal cancer (CRC). While...

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Bibliographic Details
Published in:Biosensors (Basel) 2022-02, Vol.12 (2), p.97
Main Authors: Kalligosfyri, Panagiota M, Nikou, Sofia, Karteri, Sofia, Kalofonos, Haralabos P, Bravou, Vasiliki, Kalogianni, Despina P
Format: Article
Language:English
Subjects:
Apoptosis
Biopsy
biosensor
Biosensors
Blood
Cancer
Cancer therapies
Colorectal cancer
Colorectal carcinoma
Deoxyribonucleic acid
dipstick
DNA
DNA methylation
Genotyping
gold nanoparticles
KRAS
lateral flow assay
Multiplexing
Mutation
Nanoparticles
Patients
Plasma
Precision medicine
Scientific imaging
Strip
Tumor cell lines
Tumorigenesis
Tumors
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Summary:In the era of personalized medicine, molecular profiling of patient tumors has become the standard practice, especially for patients with advanced disease. Activating point mutations of the KRAS proto-oncogene are clinically relevant for many types of cancer, including colorectal cancer (CRC). While several approaches have been developed for tumor genotyping, liquid biopsy has been gaining much attention in the clinical setting. Analysis of circulating tumor DNA for genetic alterations has been challenging, and many methodologies with both advantages and disadvantages have been developed. We here developed a gold nanoparticle-based rapid strip test that has been applied for the first time for the multiplex detection of KRAS mutations in circulating tumor DNA (ctDNA) of CRC patients. The method involved ctDNA isolation, PCR-amplification of the KRAS gene, multiplex primer extension (PEXT) reaction, and detection with a multiplex strip test. We have optimized the efficiency and specificity of the multiplex strip test in synthetic DNA targets, in colorectal cancer cell lines, in tissue samples, and in blood-derived ctDNA from patients with advanced colorectal cancer. The proposed strip test achieved rapid and easy multiplex detection (normal allele and three major single-point mutations) of the clinically relevant KRAS mutations in ctDNA in blood samples of CRC patients with high specificity and repeatability. This multiplex strip test represents a minimally invasive, rapid, low-cost, and promising diagnostic tool for the detection of clinically relevant mutations in cancer patients.
ISSN:2079-6374
2079-6374
DOI:10.3390/bios12020097