Detection of tumor progression via cell-free DNA (cfDNA) in patients with colorectal cancer.

Abstract

598 Background: Individualized therapy based on molecular stratification has become an integrative part in modern targeted treatment approaches. Here, we aimed to assess whether repeated measurements of cfDNA in patients' plasma could indicate tumor progression. Methods: We prospectively collected and stored serum from patients with colorectal cancer presenting for treatment or follow-up. KRAS and NRAS mutations were determined in the formalin fixed, paraffin embedded initial tumor material. A group of patients with confirmed KRAS or NRAS mutation, radiologically proven tumor progression and subsequent change of therapy was selected. cfDNA was extracted from samples taken at time of progression, as well as day -30 and day -60. Mutations were analyzed using a highly sensitive method based on ultradeep Next Generation Sequencing on a semiconductor-based NGS system (IonTorrent PGM), with a lower limit of detection down to 0.01% for selected mutations. Results: Twenty patients were identified to meet the inclusion criteria; 17 were KRAS mutated, three were NRAS mutated within the initial tumor material. In three patients (15%), RAS-mutation was not detectable at time of progression. While in two of them RAS-mutation had been measurable in prior samples, in one case the mutation was not detectable in cfDNA at any time. In 11 (65%) of the remaining 17 patients, RAS variant allele frequency (VAF) in the cfDNA was increased at time of progression compared to measurements at days -30 and/or -60. The median slope of increase was 3.5 (range 0.12 to 15.13). In six patients, there was no correlation between tumor progression and cfDNA; The amount of cfDNA decreased in comparison to prior measurements, either at 30 days prior to or at progression. Conclusions: In almost all cases cfDNA could be detected in the blood of patients with RAS-mutated tumors which demonstrates the high sensitivity of the method. Data suggest a trend toward an increase of cfDNA during tumor progression. However, it is not clear why deviations, especially a decrease of VAF in cfDNA occurred in some patients prior to or at tumor progression. Currently unknown tumor-biological or therapy-associated factors might contribute to this finding. Further investigation is planned.