Biotinylated Probe Isolation of Targeted Gene Region Improves Detection of T790M Epidermal Growth Factor Receptor Mutation via Peptide Nucleic Acid–Enriched Real-Time PCR

The presence of the EGFR (epidermal growth factor receptor) T790M mutation in tumor tissue or body fluids from patients treated with EGFR tyrosine kinase inhibitors may indicate the onset of resistance to treatment. It is important to identify this mutation as early as possible so that treatment can be modified accordingly or potential side effects of further treatment can be avoided. This requirement calls for high detection sensitivity. Peptide nucleic acids (PNAs) are used as PCR clamps to inhibit amplification of wild-type DNA during PCR cycling, thereby enriching for rare mutations such as T790M. We describe a modification that improves the detection limit of PNA-clamp methods by at least 20-fold.