Canine Oral Melanoma Genomic and Transcriptomic Study Defines Two Molecular Subgroups with Different Therapeutical Targets

Mucosal melanoma (MM) is a rare, aggressive clinical cancer. Despite recent advances in genetics and treatment, the prognosis of MM remains poor. Canine MM offers a relevant spontaneous and immunocompetent model to decipher the genetic bases and explore treatments for MM. We performed an integrative genomic and transcriptomic analysis of 32 canine MM samples, which identified two molecular subgroups with a different microenvironment and structural variant (SV) content. The overexpression of genes related to the microenvironment and T-cell response was associated with tumors harboring a lower content of SVs, whereas the overexpression of pigmentation-related pathways and oncogenes, such as TERT, was associated with a high SV burden. Using whole-genome sequencing, we showed that focal amplifications characterized complex chromosomal rearrangements targeting oncogenes, such as MDM2 or CDK4, and a recurrently amplified region on canine chromosome 30. We also demonstrated that the genes TRPM7, GABPB1, and SPPL2A, located in this CFA30 region, play a role in cell proliferation, and thus, may be considered as new candidate oncogenes for human MM. Our findings suggest the existence of two MM molecular subgroups that may benefit from dedicated therapies, such as immune checkpoint inhibitors or targeted therapies, for both human and veterinary medicine.

Genome-Wide Analyses for Osteosarcoma in Leonberger Dogs Reveal the CDKN2A/B Gene Locus as a Major Risk Locus

Dogs represent a unique spontaneous cancer model. Osteosarcoma (OSA) is the most common primary bone tumor in dogs (OMIA 001441-9615), and strongly resembles human forms of OSA. Several large- to giant-sized dog breeds, including the Leonberger, have a greatly increased risk of developing OSA. We performed genome-wide association analysis with high-density imputed SNP genotype data from 273 Leonberger cases with a median age of 8.1 [3.1–13.5] years and 365 controls older than eight years. This analysis revealed significant associations at the CDKN2A/B gene locus on canine chromosome 11, mirroring previous findings in other dog breeds, such as the greyhound, that also show an elevated risk for OSA. Heritability (h2SNP) was determined to be 20.6% (SE = 0.08; p-value = 5.7 × 10−4) based on a breed prevalence of 20%. The 2563 SNPs across the genome accounted for nearly all the h2SNP of OSA, with 2183 SNPs of small effect, 316 SNPs of moderate effect, and 64 SNPs of large effect. As with many other cancers it is likely that regulatory, non-coding variants underlie the increased risk for cancer development. Our findings confirm a complex genetic basis of OSA, moderate heritability, and the crucial role of the CDKN2A/B locus leading to strong cancer predisposition in dogs. It will ultimately be interesting to study and compare the known genetic loci associated with canine OSA in human OSA.

Whole Genome Sequencing Reveals Multiple Linked Genetic Variants on Canine Chromosome 12 Associated with Risk for Symmetrical Lupoid Onychodystrophy (SLO) in the Bearded Collie

In dogs, symmetrical lupoid onychodystrophy (SLO) results in nail loss and an abnormal regrowth of the claws. In Bearded Collies, an autoimmune nature has been suggested because certain dog leukocyte antigen (DLA) class II haplotypes are associated with the condition. A genome-wide association study of the Bearded Collie revealed two regions of association that conferred risk for disease: one on canine chromosome (CFA) 12 that encompasses the DLA genes, and one on CFA17. Case-control association was employed on whole genome sequencing data to uncover putative causative variants in SLO within the CFA12 and CFA17 associated regions. Genotype imputation was then employed to refine variants of interest. Although no SLO-associated protein-coding variants were identified on CFA17, multiple variants, many with predicted damaging effects, were identified within potential candidate genes on CFA12. Furthermore, many potentially damaging alleles were fully correlated with the presence of DLA class II risk haplotypes for SLO, suggesting that the variants may reflect DLA class II haplotype association with disease or vice versa. Strong linkage disequilibrium in the region precluded the ability to isolate and assess the individual or combined effect of variants on disease development. Nonetheless, all were predictive of risk for SLO and, with judicious assessment, their application in selective breeding may prove useful to reduce the incidence of SLO in the breed.

Identification of Aneuploidy in Dogs Screened by a SNP Microarray

Microarray analysis is an efficient approach for screening and identifying cytogenetic imbalances in humans. SNP arrays, in particular, are a powerful way to identify copy number gains and losses representing aneuploidy and aneusomy, but moreover, allow for the direct assessment of individual genotypes in known disease loci. Using these approaches, trisomies, monosomies and mosaicism of whole chromosomes have been identified in human microarray studies. For canines, this approach is not widely used in clinical laboratory diagnostic practice. In our laboratory, we have implemented the use of a propriety SNP array that represents approximately 650,000 loci across the domestic dog genome. During the validation of this microarray prior to clinical use, we identified three cases of aneuploidy after screening 2,053 dogs of various breeds including monosomy X, trisomy X and an apparent, mosaic trisomy of canine chromosome 38 (CFA 38). This study represents the first use of microarrays for copy number evaluation to identify cytogenetic anomalies in canines. As microarray analysis becomes more routine in canine genetic testing, more cases of chromosome aneuploidy are likely to be uncovered.

Variants within the SP110 nuclear body protein modify risk of canine degenerative myelopathy

Canine degenerative myelopathy (DM) is a naturally occurring neurodegenerative disease with similarities to some forms of amyotrophic lateral sclerosis (ALS). Most dogs that develop DM are homozygous for a common superoxide dismutase 1 gene (SOD1) mutation. However, not all dogs homozygous for this mutation develop disease. We performed a genome-wide association analysis in the Pembroke Welsh Corgi (PWC) breed comparing DM-affected and -unaffected dogs homozygous for the SOD1 mutation. The analysis revealed a modifier locus on canine chromosome 25. A haplotype within the SP110 nuclear body protein (SP110) was present in 40% of affected compared with 4% of unaffected dogs (P = 1.5 × 10−5), and was associated with increased probability of developing DM (P = 4.8 × 10−6) and earlier onset of disease (P = 1.7 × 10−5). SP110 is a nuclear body protein involved in the regulation of gene transcription. Our findings suggest that variations in SP110-mediated gene transcription may underlie, at least in part, the variability in risk for developing DM among PWCs that are homozygous for the disease-related SOD1 mutation. Further studies are warranted to clarify the effect of this modifier across dog breeds.

Relevance of chromosome 13 aberrations in canine tumours

SummaryFor human tumours there are many reports documenting the correlation between chromosome aberrations and tumour entities. Due to the complex canine karyotypic pattern (78 chromosomes), cytogenetic studies of tumours of the dog are rare. However, the reports in the literature show, that canine chromosome 13 (CFA 13) is predominantly involved in chromosomal changes. Interestingly, CFA 13 shows high homology to regions on the human chromosomes 4 (HSA 4) and 8 (HSA 8), which harbour the proto-oncogenes c-KIT and c-MYC. Both of these genes are involved in the development and progression of some human and canine tumour diseases.

Progressive rod-cone degeneration (PRCD) in selected dog breeds and variability in its phenotypic expression

Progressive rod-cone degeneration (PRCD) is a late onset autosomal photoreceptor degeneration found in canines. PRCD in canines is homologous to one form of retinitis pigmentosa (RP) found in humans and displays phenotypic similarity as well as having the identical causative mutation. The PRCD gene was mapped to the centromeric region of canine chromosome 9 (CFA9). We report here a population study of 699 dogs of the following breeds and the following frequencies of the disease-causing mutation: American Cocker Spaniel (0.09), English Cocker Spaniel (0.34), English Springer Spaniel (0.00), Welsh Springer Spaniel (0.00), Flat Coated Retriever (0.00), Golden Retriever (0.00), Chesapeake Bay Retriever (0.14), Nova Scotia Duck Tolling Retriever (0.44), Labrador Retriever (0.07), Poodle Toy (0.45), Poodle Miniature (0.20), Poodle Medium (0.05), Poodle Standard (0.00), Portuguese Water Dog (0.33), Chinese Crested Dog (0.02), Shipperke (0.06), and Australian Cattle Dog (0.00). The disease results in complete blindness in the affected individual in almost every case. The time of onset and disease progression varies between dog breeds as well as between individuals. A modifier gene is likely to segregate in genomic proximity to the PRCD gene and may influence phenotypic expression.