Clinical utility of liquid biopsy for monitoring patients with NRAS‐mutant medium‐to‐giant congenital melanocytic nevi

Dear Editor, Congenital melanocytic nevi (CMN) are benign melanocytic skin tumours that are classified according to the projected adult size (PAS), the categorization developed by Krengel et al., and the “6B” distribution patterns. Patients with multiple medium CMN (PAS 1.5– 20 cm), large CMN (PAS 20– 40 cm) and giant CMN (PAS >40 cm) have an increased lifetime risk of melanoma (10– 15%) and neurocutaneous melanocytosis (NMC) (12%), which may be associated with central nervous system (CNS) malformations, including the Dandy– Walker malformation (DWM) complex. While monitoring driver mutations in cellfree DNA (cfDNA) is a relevant strategy for managing melanoma patients, its clinical utility in CMN patients remains unknown. Here, we aimed to analyse the clinical utility of the analysis of NRAS mutations in cellfree DNA (cfNRAS) for monitoring NRASmutant mediumtogiant CMN patients (~80% of cases). After institutional approval and written informed consent from patients or their legal tutor, we collected 21 plasma samples from 14 NRASmutant mediumtogiant CMN patients using CellFree DNA BCT tubes (Streck Inc) (Table 1). cfNRAS mutation screening was performed using the droplet digital PCR System and two commercial assays (BioRad Laboratories). All samples were run in triplicate and a positive result was considered when mutationpositive droplets were detected in ≥2 replicates. Results were reported as the variant allele frequency (VAF). Patients 1 to 10 presented with CMN of different PAS and no CMNassociated comorbidities (Table 1). No cfNRAS was detected in any of their samples (n = 12), suggesting that benign melanocytes of CMN do not shed cfNRAS into the bloodstream, regardless of the lesion PAS. This result is in line with a previous observation in healthy individuals, in which the presence of multiple acquired melanocytic nevi did not promote the detection of BRAF mutations in cfDNA. Patient 11 had a giant CMN and asymptomatic radiological DWM complex. We analysed two longitudinal plasma samples and no cfNRAS was detected in either sample (Table 1). Patient 12 had multiple medium CMN and symptomatic NCM. At age 5, he was diagnosed with CNS melanoma and died 3 months after. We obtained a plasma sample during disease progression but no cfNRAS was detected in it (Table 1). These two cases show that plasma cfDNA screening has limited value to detect CNS abnormalities, probably due to the presence of the blood– brain barrier. Cerebrospinal f luid might be a better option for minimally invasive diagnosis and treatment monitoring in patients with highly suspected or confirmed tumours within the CNS. Patient 13 had a giant CMN, severe NCM, hydrocephalus, DWM complex and congenital adrenal hyperplasia. We detected cfNRAS in a plasma sample obtained when she was under laser hairremoval treatment of the CMN (VAF = 12.9%). She had no suspicious signs of cutaneous or extracutaneous malignancy at the time of plasma collection or after 2 years of followup. VAF turned to 0% in a sample obtained 18 months after the last laser session (Table 1). This data points to the laser as a likely modifier of melanocyte turnover and, in turn, a potential promoter of cfNRAS shedding. Other treatments, such as tissue expansion, curettage and dermabrasion, may also affect melanocyte turnover. Therefore, treatment history details should be reported when liquid biopsy strategies are applied in CMN patients. Last, Patient 14 had a nodal metastatic melanoma associated with a Spilustype large CMN. While no cfNRAS was detected when he was free of disease, progressively increasing levels of cfNRAS were detected during disease progression, with a clear correlation between the tumour burden and the VAF (0.6% to 82% just before dying) (Figure 1, Table 1). This preliminary data suggests that cfNRAS might be a potential biomarker for early detection of melanoma progression outside the CNS and subsequent monitoring in NRASmutant mediumtogiant CMN patients. An expansion to a larger number of patients would be useful to confirm the clinical utility of liquid biopsies in the context of this rare disease. Received: 7 May 2022 | Accepted: 1 September 2022