Comprehensive Genetic Landscape of Uveal Melanoma by Whole-Genome Sequencing

Abstract

Uveal melanoma (UM) is a rare intraocular tumor that, similar to cutaneous melanoma, originates from melanocytes. To gain insights into its genetics, we performed whole-genome sequencing at very deep coverage of tumor-control pairs in 33 samples (24 primary and 9 metastases). Genome-wide, the number of coding mutations was rather low (only 17 variants per tumor on average; range 7-28), thus radically different from cutaneous melanoma, where hundreds of exonic DNA insults are usually detected. Furthermore, no UV light-induced mutational signature was identified. Recurrent coding mutations were found in the known UM drivers GNAQ, GNA11, BAP1, EIF1AX, and SF3B1. Other genes, i.e., TP53BP1, CSMD1, TTC28, DLK2, and KTN1, were also found to harbor somatic mutations in more than one individual, possibly indicating a previously undescribed association with UM pathogenesis. De novo assembly of unmatched reads from non-coding DNA revealed peculiar copy-number variations defining specific UM subtypes, which in turn could be associated with metastatic transformation. Mutational-driven comparison with other tumor types showed that UM is very similar to pediatric tumors, characterized by very few somatic insults and, possibly, important epigenetic changes. Through the analysis of whole-genome sequencing data, our findings shed new light on the molecular genetics of uveal melanoma, delineating it as an atypical tumor of the adult for which somatic events other than mutations in exonic DNA shape its genetic landscape and define its metastatic potential.

Figure 1

Unsupervised Hierarchical Clustering and Global Genetic Landscape of All Tumors Analyzed in This Study Sample IDs are indicated on the right. CNV events are depicted in blue (copy losses) or in shades of red (copy gains) and ploidy is indicated in the legend provided at the top. SNVs in genes found to carry mutations in six or more individuals are shown on the left, with mutation classes provided within gray or blue boxes. Clustering identifies four classes—A, B, C, and D (see text)—indicated within the dendrogram.

Figure 2

Inferred Somatic Events Defining Tumor Classes, as Identified by Clustering Colors are the same as those shown in Figure 1. All steps determining branching are statistically significant.

Figure 3

Genetic Features in Primary and Metastatic Tumors (A) Overview of all major somatic events with respect to PUMs and MUMs. Each circle indicates a specific genetic event; its center corresponds to the percentage of samples carrying this feature in PUMs versus MUMs, whereas its diameter indicates the total number of such samples. Asterisks indicate statistical significance. The gray area depicts the surface of the plot for which there is an enrichment in MUMs. (B–E) Boxplots of different types of genetic alterations, at the genome-wide scale.

Figure 4

Landscape of Genetic Alterations Involving BAP1 and Immunohistochemistry of BAP1 Protein (A) Samples are ordered with respect to absence/presence of BAP1 nuclear localization, indicating loss or preservation of protein function, respectively. In general, loss of BAP1 function correlates with presence of a somatic SNV and loss of heterozygosity. (B and C) Representative micrographs of paraffin-embedded UM samples showing absence (B) and presence (C) of BAP1 protein. Magnification: 252×.

Figure 5

Mutational Signature of Our Samples, for SNSs, Genome-wide (A) Main graph: comparison of mutational load of the UM samples studied with respect to two melanomas of the conjunctiva (CM), sequenced and processed according to the same methods. The number of mutations is radically different in UM versus CM. Inset: mutational spectrum of each UM sample, in percentage, showing a relatively homogeneous pattern. (B) Results of the analysis of mutational events according to the methods and the classification proposed by Alexandrov et al., Three main signatures are detected in our samples, evocative of Alexandrov’s signatures 12/16, 1B, and 6. The different peaks indicate specific genetic contexts of the altered nucleotide and are ordered according to the original article.

Figure 6

Analysis of the Mutational Spectrum in Our Samples versus Other Cancer Types, in Coding Regions Only (A) The spectrum from UM is dissimilar from those from cutaneous and conjunctival melanomas, which are dominated by UV light-induced events (C to T transitions) and is conversely closer to that of thyroid and renal papillary cancer. (B) Principal-component analysis (PCA) of the same data, showing the relatedness of UM with a few cancers but, again, not with other melanomas. Dimensions of the PCA are indicated by the arrowed axes. Primary data other than UM are from previously published sources., Cancer types are indicated by the following abbreviations. aLung: lung, adenocarcinoma; scLung: lung, squamous cell; pRenal: renal, papillary; ccRenal: renal, clear cell; GBM: glioblastoma multiforme; AML: acute myeloid leukemia; Cut_mel: cutaneous melanoma; CM: conjunctival melanoma.

Figure 7

Circos Plot of All Somatic Interchromosomal Events, in All UM Samples Red lines indicate events involving the same chromosomal regions in more than two individuals.