Dynamic ctDNA Mutational Complexity in Patients with Melanoma Receiving Immunotherapy.

BACKGROUND: Circulating tumour DNA (ctDNA) analysis promises to improve the clinical care of people with cancer, address health inequities and guide translational research. This observational cohort study used ctDNA to follow 29 patients with advanced-stage cutaneous melanoma through multiple cycles of immunotherapy. METHOD: A melanoma-specific ctDNA next-generation sequencing (NGS) panel, droplet digital polymerase chain reaction (ddPCR) and mass spectrometry analysis were used to identify ctDNA mutations in longitudinal blood plasma samples from Aotearoa New Zealand (NZ) patients receiving immunotherapy for melanoma. These technologies were used in conjunction to identify the breadth and complexity of tumour genomic information that ctDNA analysis can reliably report. RESULTS: During the course of immunotherapy treatment, a high level of dynamic mutational complexity was identified in blood plasma, including multiple BRAF mutations in the same patient, clinically relevant BRAF mutations emerging through therapy and co-occurring sub-clonal BRAF and NRAS mutations. The technical validity of this ctDNA analysis was supported by high sample analysis-reanalysis concordance, as well as concordance between different ctDNA measurement technologies. In addition, we observed > 90% concordance in the detection of ctDNA when using cell-stabilising collection tubes followed by 7-day delayed processing, compared with standard EDTA blood collection protocols with rapid processing. We also found that the undetectability of ctDNA at a proportion of treatment cycles was associated with durable clinical benefit (DCB). CONCLUSION: We found that multiple ctDNA processing and analysis methods consistently identified complex longitudinal patterns of clinically relevant mutations, adding support for expanded clinical trials of this technology in a variety of oncology settings.