Detection of Epidermal Growth Factor Receptor (EGFR) Mutations in Circulating Tumor DNA during EGFR-Tyrosine Kinase Inhibitor (EGFR-TKI) Treatment

Background

Knowledge of EGFR mutation status is a crucial step to select the best treatment in first line as well as second line setting after the development of acquired resistance. Obtaining Tumor tissue or cytology samples are not always available, and rebiopsy is not easy to perform. Recently, studies showed that circulating tumor DNA can be used as a suitable substitute for mutation analysis. We compared the sensitivity of EGFR mutation detection techniques from tumor tissue and circulating tumor DNA in patients being treated with EGFR-TKIs.

Methods

We collected plasma samples before treatment with EGFR-TKI and after acquired resistance in 11 patients with activating EGFR mutations (5 cases with exon 19 deletion and 6 cases of L858R) in DNA extracted from paraffin-embedded tissues using PNA Clamp EGFR Mutation Detection kit (Panagen, Korea). The extraction of circulating tumor DNA (CtDNA) from plasma was performed using QIAamp circulating nucleic acid kit (Qiagen, Germany). EGFR mutation analysis for CtDNA was performed by pyrosequencing and PANA Mutyper R EGFR kit (Panagene, Korea).

Results

The EGFR mutation detection sensitivity of pyrosequencing from CtDNA was 33.3%, and that of PANA Mutyper was 72.7%. The degree of agreements between tissue DNA and CtDNA were better in PANA Mutyper (k=0.429, p=0.033) than pyrosequencing (k=0.194, p=0.087). After the development of acquired resistance, same EGFR mutations were detected in 63.6% by PANA Mutyper from CtDNA. One of them showed newly developed T790M mutation.

Conclusion

PANA Mutyper test were better than pyrosequencing in the sensitivity and the strength of agreement between CtDNA and tissue DNA. The detection sensitivity of T790M resistance mutation from CtDNA should be validated with more sensitive technique.