Plasma epidermal growth factor receptor mutation testing with a chip-based digital PCR system in patients with advanced non-small cell lung cancer.

OBJECTIVES: Epidermal growth factor receptor (EGFR) mutation testing is a companion diagnostic to determine eligibility for treatment with EGFR tyrosine kinase inhibitors (EGFR-TKIs) in non-small cell lung cancer (NSCLC). Recently, plasma-based EGFR testing by digital polymerase chain reaction (dPCR), which enables accurate quantification of target DNA, has shown promise as a minimally invasive diagnostic. Here, we aimed to evaluate the accuracy of a plasma-based EGFR mutation test developed using chip-based dPCR-based detection of 3 EGFR mutations (exon 19 deletions, L858R in exon 21, and T790M in exon 20). MATERIALS AND METHODS: Forty-nine patients with NSCLC harboring EGFR-activating mutations were enrolled, and circulating free DNAs (cfDNAs) were extracted from the plasma of 21 and 28 patients before treatment and after progression following EGFR-TKI treatment, respectively. RESULTS: Using reference genomic DNA containing each mutation, the detection limit of each assay was determined to be 0.1%. The sensitivity and specificity of detecting exon 19 deletions and L858R mutations, calculated by comparing the mutation status in the corresponding tumors, were 70.6% and 93.3%, and 66.7% and 100%, respectively, showing similar results compared with previous studies. T790M was detected in 43% of 28 cfDNAs after progression with EGFR-TKI treatment, but in no cfDNAs before the start of the treatment. CONCLUSION: This chip-based dPCR assay can facilitate detection of EGFR mutations in cfDNA as a minimally invasive method in clinical settings.

Metabolic profiling of gemcitabine- and paclitaxel-treated immortalized human pancreatic cell lines with K-RASG12D.

The mechanisms of action of gemcitabine (GEM) and paclitaxel (PTX) have been well investigated, and shown to be the inhibition of DNA polymerase and polymerization of tubulin, respectively. Meanwhile, genomic research has revealed that mutations in the K-RAS oncogene occur in over 90% of pancreatic cancer. Oncogenic alteration rewires alternative metabolic pathways to satisfy the demands of growth. The K-RAS oncogene also has been shown to upregulate glycolysis and glutaminolysis. However, it is still unclear whether K-RAS independently plays a central role in controlling tumor metabolism. Here, we conducted a metabolomic analysis of a simple oncogenic K-RAS cell line model constructed using human telomerase catalytic subunit-immortalized human pancreatic epithelial nestin-expressing cell lines with and without K-RASG12D. We also investigated the effect of GEM and PTX on these cells. As a result, it was shown in the cell with K-RASG12D that the level of lactate was increased and glutamic acid, glutamine, and aspartic acid levels were decreased. In the nucleotide metabolism, GEM-treated cells showed metabolic changes, whereas these phenomena were not observed in PTX-treated cells. In conclusion, it was suggested that K-RASG12D independently modified tumor metabolism and the difference between GEM and PTX in the nucleotide metabolism was revealed.