Back close

An allele-specific quantitative PCR strategy for detection of EGFR T790M mutations by liquid biopsy in non-small cell lung cancer patients

Publication Type : Journal Article

Publisher : Elsevier BV

Source : International Journal of Biological Macromolecules

Url : https://doi.org/10.1016/j.ijbiomac.2025.144447

Keywords : Circulating free nucleic acid, NSCLC, Tyrosine kinase inhibitors

Campus : Kochi

School : School of Pharmacy

Year : 2025

Abstract : Detection of mutations conferring resistance to tyrosine kinase inhibitors (TKIs) in epidermal growth factor receptor (EGFR), such as T790M, is critical for guiding targeted therapies in non-small cell lung cancer (NSCLC) patients. However, limitations in accessing tumor tissue have given rise to liquid biopsy approaches necessitating ultrasensitive techniques to identify tumor mutations in a large background of wild-type alleles. We developed a sensitive and cost-effective technique utilizing a 2′3’-dideoxy nucleotide (ddNTP) blocker that selectively blocks wild-type allele amplification, allowing specific detection of mutant alleles in circulating free DNA (cfDNA) from plasma. EGFR genomic sequences encompassing T790M were cloned into pMD20-T plasmids and titrated in a background of genomic DNA to test the sensitivity. The assay detected 10 copies of the mutant allele in the presence of 10,000 copies of wild-type alleles, indicating a sensitivity of 0.001 %. Among 74 NSCLC patients, 34 received TKI treatment and were screened for the T790M mutation. The current method would be a sensitive and cost-effective liquid biopsy tool for detecting EGFR T790M mutations in NSCLC cfDNA samples, with strong potential for identifying actionable mutations for targeted therapy, especially in low-resource settings.

Cite this Research Publication : Gayathri Baburaj, Preetiparna Parida, Karthik S. Udupa, Ananth Pai, Rama Rao Damerla, Mahadev Rao, An allele-specific quantitative PCR strategy for detection of EGFR T790M mutations by liquid biopsy in non-small cell lung cancer patients, International Journal of Biological Macromolecules, Elsevier BV, 2025, https://doi.org/10.1016/j.ijbiomac.2025.144447

Admissions Apply Now