68P - Comparison of methods for circulating cell-free DNA isolation using blood from cancer patients. Impact on biomarker testing

Abstract

The implementation of liquid biopsy for biomarker testing and response to treatment monitoring requires the development of automated methods for circulating free DNA (cfDNA) isolation. Nevertheless, the extraction stage is critical to ensure reliable results. The present study compares the MagNA Pure Compact Nucleic Acid Isolation (MPC) Kit I and Maxwell® RSC ccfDNA Plasma Kit (MR) and the later with QIAamp Circulating Nucleid Acid (QCNA) kit using 57 serum samples from cancer patients treated at the Medical Oncology Department at Puerta de Hierro Teaching Hospital in Madrid, Spain. Firstly, we observed that MPC method significantly extracted less cfDNA than MR (P < 0.0001). However, there were no significant differences in extraction yields of QCNA and MR kits. cfDNA isolation yield was also associated with tumor stage but not with tumor location. Secondly, an oligonucleosomal DNA ladder pattern was observed in 88% of the samples and significant differences in the recovery of mono, di- and trinucleosomes DNA fragments were observed between MPC and MR methodologies. Finally, tumor mutation quantification on cfDNA was performed on 38 paired samples using digital PCR. Mutant allele fractions between paired samples were not significantly different. Methods for isolation of cfDNA can affect DNA yield and molecular weight fractions recovery. These observations should be taken into account for cfDNA analysis in routine clinical practice in cancer patients.