Abstract
The presence of cell-free DNA (cfDNA) in plasma from non-small cell lung cancer (NSCLC) patients demonstrates promising diagnostic implications as the minimally-invasive liquid biopsy. Recently, we showed promising but clinically not satisfactory sensitivity/specificity of plasma DNA quantification for NSCLC detection. Mean cfDNA level in 66 NSCLC patients (21.5 ng/ml) was significantly higher than in 40 healthy controls (4.5 ng/ml; p=0.000), but did not differ from values observed in 15 patients with non-malignant tumors (23.4 ng/ml; hamartoma, fibrosis, granuloma). Similarly, mean cfDNA integrity index significantly differed NSCLC (4.0) from healthy controls (1.0; p=0.000), but not from non-malignant group (4.0).
To improve the diagnostic power of cfDNA analysis, we introduced a simultaneous methylation profiling of 21 tumor suppressor genes (TSGs) in plasma cfDNA from NSCLC versus non-malignant patients using MS-MLPA assay. 25/32 (78%) NSCLC and 8/8 (100%) hamartoma cfDNA samples presented at least one TSG methylation. APC, ATM, DAPK1, HIC1, MLH1 and RARβ were the most frequently methylated genes in NSCLC, while MLH1, TIMP3, TP73 – in hamartoma patients.
We demonstrated that optimized MS-MLPA assay allows detection multiple methylated TSGs in plasma cfDNA. The MS-MLPA showed good performance in samples with diverse cfDNA concentrations suggesting that methylation detection rate depends on the methylated DNA content in a sample. The TSG methylation in hamartoma patients may indicate an increased risk of lung cancer. The groups are to be enlarged, TSG panel extended up to 37 genes and paired plasma-tumor tissue analyzed.
- © 2014 ERS
