Combined mutation and copy-number variation detection by targeted next-generation sequencing in uveal melanoma



Smit, Kyra N, van Poppelen, Natasha M, Vaarwater, Jolanda, Verdijk, Robert, van Marion, Ronald, Kalirai, Helen ORCID: 0000-0002-4440-2576, Coupland, Sarah E ORCID: 0000-0002-1464-2069, Thornton, Sophie ORCID: 0000-0001-7693-7279, Farquhar, Neil, Dubbink, Hendrikus-Jan
et al (show 3 more authors) (2018) Combined mutation and copy-number variation detection by targeted next-generation sequencing in uveal melanoma. MODERN PATHOLOGY, 31 (5). pp. 763-771.

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Abstract

Uveal melanoma is a highly aggressive cancer of the eye, in which nearly 50% of the patients die from metastasis. It is the most common type of primary eye cancer in adults. Chromosome and mutation status have been shown to correlate with the disease-free survival. Loss of chromosome 3 and inactivating mutations in BAP1, which is located on chromosome 3, are strongly associated with 'high-risk' tumors that metastasize early. Other genes often involved in uveal melanoma are SF3B1 and EIF1AX, which are found to be mutated in intermediate- and low-risk tumors, respectively. To obtain genetic information of all genes in one test, we developed a targeted sequencing method that can detect mutations in uveal melanoma genes and chromosomal anomalies in chromosome 1, 3, and 8. With as little as 10 ng DNA, we obtained enough coverage on all genes to detect mutations, such as substitutions, deletions, and insertions. These results were validated with Sanger sequencing in 28 samples. In >90% of the cases, the BAP1 mutation status corresponded to the BAP1 immunohistochemistry. The results obtained in the Ion Torrent single-nucleotide polymorphism assay were confirmed with several other techniques, such as fluorescence in situ hybridization, multiplex ligation-dependent probe amplification, and Illumina SNP array. By validating our assay in 27 formalin-fixed paraffin-embedded and 43 fresh uveal melanomas, we show that mutations and chromosome status can reliably be obtained using targeted next-generation sequencing. Implementing this technique as a diagnostic pathology application for uveal melanoma will allow prediction of the patients' metastatic risk and potentially assess eligibility for new therapies.

Item Type: Article
Uncontrolled Keywords: Chromosomes, Human, Pair 1, Chromosomes, Human, Pair 3, Humans, Melanoma, Uveal Neoplasms, Ubiquitin Thiolesterase, Tumor Suppressor Proteins, Phosphoproteins, Eukaryotic Initiation Factor-1, DNA, Neoplasm, Prognosis, Disease-Free Survival, In Situ Hybridization, Fluorescence, Mutation, Polymorphism, Single Nucleotide, High-Throughput Nucleotide Sequencing, RNA Splicing Factors
Depositing User: Symplectic Admin
Date Deposited: 16 Apr 2019 14:43
Last Modified: 19 Jan 2023 06:33
DOI: 10.1038/modpathol.2017.187
Related URLs:
URI: https://livrepository.liverpool.ac.uk/id/eprint/3021190