Circulating Tumor DNA as a Marker for Treatment Response in Metastatic Melanoma Patients Using Next-Generation Sequencing—A Prospective Feasibility Study

SIMPLE SUMMARY: Despite the improvement of the prognosis of metastatic melanoma patients through the implementation of targeted and immunotherapies, there is a need to identify biomarkers to predict and monitor treatment response. Therefore, we performed sequencing of paired melanoma tissue biopsies and circulating tumor DNA (ctDNA) from 149 plasma samples using two custom next-generation sequencing (NGS) AmpliSeq™ HD panels to determine the level of concordance. We aimed to evaluate whether ctDNA analysis with NGS could predict and monitor treatment response in a cohort of metastatic melanoma patients; NGS allowed for a comprehensive analysis of cancer-associated mutations in serial plasma samples with high sensitivity. Although a trend could be seen that mutant allele frequency levels over time correlated with or even preceded radiological response to treatment, this finding was not statistically significant in our cohort. Our study demonstrates that NGS gene panels might be useful for treatment monitoring with ctDNA in melanoma patients. ABSTRACT: We prospectively performed a longitudinal analysis of circulating tumor DNA (ctDNA) from 149 plasma samples and CT scans in Stage III and IV metastatic melanoma patients (n = 20) treated with targeted agents or immunotherapy using two custom next-generation sequencing (NGS) Ion AmpliSeq™ HD panels including 60 and 81 amplicons in 18 genes, respectively. Concordance of matching cancer-associated mutations in tissue and plasma was 73.3%. Mutant allele frequency (MAF) levels showed a range from 0.04% to 28.7%, well detectable with NGS technologies utilizing single molecule tagging like the AmpliSeq™ HD workflow. Median followup time of the tissue and/or plasma positive cohort (n = 15) was 24.6 months and median progression-free survival (PFS) was 7.8 months. Higher MAF ≥ 1% at baseline was not significantly associated with a risk of progression (Odds Ratio = 0.15; p = 0.155). Although a trend could be seen, MAF levels did not differ significantly over time between patients with and without a PFS event (p = 0.745). Depending on the cell-free DNA amount, NGS achieved a sensitivity down to 0.1% MAF and allowed for parallel analysis of multiple mutations and previously unknown mutations. Our study indicates that NGS gene panels could be useful for monitoring disease burden during therapy with ctDNA in melanoma patients.