Canine malignant melanoma is a common cancer with a high mortality rate. Although previous studies have evaluated various aspects of this tumour, the exact mechanism of tumourigenesis remains unknown. Epigenetic mechanisms, such as DNA methylation, have recently gained attention as aetiological factors for neoplasia in humans. This study aimed to analyse genome-wide DNA methylation patterns in canine malignant melanoma based on next-generation sequencing data.

A total of 76,213 CpG sites, including 29,482 sites in CpG islands (CGIs), were analysed using next-generation sequencing of methylation-specific signatures, obtained by sequential digestion with enzymes, to compare normal oral mucosal samples from four healthy dogs, four canine melanoma cell lines (3 oral cavity and 1 skin), and five clinical samples of oral canine melanoma.

Malignant melanoma showed increased methylation at thousands of normally unmethylated CpG sites in CGIs and decreased methylation at normally methylated CpG sites in non-CGIs. Interestingly, the promoter regions of 81–393 genes were hypermethylated; 23 of these genes were present in all melanoma cell lines and melanoma clinical samples. Among these 23 genes, six genes with “sequence-specific DNA binding” annotation were significantly enriched, including three Homeobox genes—HMX2, TLX2, and HOXA9—that may be involved in the tumourigenesis of canine malignant melanoma.

This study revealed widespread alterations in DNA methylation and a large number of hypermethylated genes in canine malignant melanoma.

犬恶性黑色素瘤是死亡率高的常见癌症。尽管先前的研究已经评估了该肿瘤的各个方面，但肿瘤发生的确切机制仍然未知。表观遗传机制，例如DNA甲基化，最近已引起人们的关注，成为人类肿瘤形成的病因。这项研究旨在根据下一代测序数据分析犬恶性黑色素瘤的全基因组DNA甲基化模式。

使用下一代甲基化特异性标记序列（通过酶顺序消化获得），对总共76,213个CpG位点（包括CpG岛（CGI）中的29,482个位点）进行了分析，以比较来自四只健康狗，四只犬的正常口腔粘膜样本黑色素瘤细胞系（3个口腔和1个皮肤），以及五个犬类黑色素瘤临床样品。

恶性黑色素瘤在CGI的数千个正常未甲基化的CpG位点显示甲基化增加，而在非CGI的正常甲基化的CpG位点显示甲基化降低。有趣的是，81-393个基因的启动子区域被甲基化。这些基因中的23个存在于所有黑色素瘤细胞系和黑色素瘤临床样品中。在这23个基因中，有6个带有“序列特异性DNA结合”注释的基因得到了显着富集，包括3个Homeobox基因（HMX2，TLX2和HOXA9），它们可能与犬恶性黑色素瘤的肿瘤发生有关。

这项研究揭示了犬恶性黑色素瘤中DNA甲基化的广泛变化和大量的高甲基化基因。