Towards Personalized Medicine in Melanoma: Implementation of a Clinical Next-Generation Sequencing Panel.

de Unamuno Bustos B1, Murria Estal R2, Pérez Simó G2, de Juan Jimenez I2, Escutia Muñoz B1, Rodríguez Serna M1, Alegre de Miquel V3, Llavador Ros M4, Ballester Sánchez R5, Nagore Enguídanos E6, Palanca Suela S7, Botella Estrada R1.

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Abstract

Molecular diagnostics are increasingly performed routinely in the diagnosis and management of patients with melanoma due to the development of novel therapies that target specific genetic mutations. The development of next-generation sequencing (NGS) technologies has enabled to sequence multiple cancer-driving genes in a single assay, with improved sensitivity in mutation detection. The main objective of this study was the design and implementation of a melanoma-specific sequencing panel, and the identification of the spectrum of somatic mutations in a series of primary melanoma samples. A custom panel was designed to cover the coding regions of 35 melanoma-related genes. Panel average coverage was 2,575.5 reads per amplicon, with 92,8% of targeted bases covered ≥500×. Deep coverage enabled sensitive discovery of mutations in as low as 0.5% mutant allele frequency. Eighty-five percent (85/100) of the melanomas had at least one somatic mutation. The most prevalent mutated genes were BRAF (50%;50/199), NRAS (15%;15/100), PREX2 (14%;14/100), GRIN2A (13%;13/100), and ERBB4 (12%;12/100). Turn-around-time and costs for NGS-based analysis was reduced in comparison to conventional molecular approaches. The results of this study demonstrate the cost-effectiveness and feasibility of a custom-designed targeted NGS panel, and suggest the implementation of targeted NGS into daily routine practice.