Determination of serum miRNAs biomarkers of lung cancer by quantitative PCR (qPCR) and by droplet digital PCR (ddPCR). Determinazione dei livelli sierici dei miRNA come biomarcatori del cancro al polmone mediante PCR quantitativa (qPCR) e droplet digital PCR (ddPCR).

Lung cancer (LC) is often diagnosed at an advanced stage, when it is not radically curable. Minimally invasive methods allowing identification of asymptomatic subjects with early LC are urgently needed and selected microRNAs (miRNAs) have been suggested as circulating cell-free biomarkers of LC. We compared the performance of two methods, namely qPCR (both absolute and relative) and droplet digital PCR (ddPCR), in accurately measuring the levels of circulating miRNAs of interest for LC. It was found that ddPCR is more precise and provides higher throughput of analysis than standard qPCR, at a similar cost-per-sample; moreover, ddPCR does not rely on external calibrators or reference genes. Following systematic review of the pertinent literature, eight miRNAs of interest for LC were identified and measured by ddPCR in the serum of 85 patients with early LC (stage I and II) and 83 controls. Four miRNAs (let-7a, miR-210, miR-320a, miR-221) showed significantly different serum level in LC patients and in controls. For each of these miRNAs, the Receiver Operating Characteristic curve was constructed and the Area Under the Curve showed fair accuracy in identifying LC cases (about 0.7 for each miRNA). In conclusion, ddPCR was a robust method for absolute quantification of miRNAs of interest for LC. Testing the combination of the above four miRNAs in the serum may help identifying subjects with early LC.