Whole exome sequencing analysis of canine urothelial carcinomas without BRAF V595E mutation: Short in-frame deletions in BRAF and MAP2K1 suggest alternative mechanisms for MAPK pathway disruption.

Molecular profiling studies have shown that 85% of canine urothelial carcinomas (UC) harbor an activating BRAF V595E mutation, which is orthologous to the V600E variant found in several human cancer subtypes. In dogs, this mutation provides both a powerful diagnostic marker and a potential therapeutic target; however, due to their relative infrequency, the remaining 15% of cases remain understudied at the molecular level. We performed whole exome sequencing analysis of 28 canine urine sediments exhibiting the characteristic DNA copy number signatures of canine UC, in which the BRAF V595E mutation was undetected (UDV595E specimens). Among these we identified 13 specimens (46%) harboring short in-frame deletions within either BRAF exon 12 (7/28 cases) or MAP2K1 exons 2 or 3 (6/28 cases). Orthologous variants occur in several human cancer subtypes and confer structural changes to the protein product that are predictive of response to different classes of small molecule MAPK pathway inhibitors. DNA damage response and repair genes, and chromatin modifiers were also recurrently mutated in UDV595E specimens, as were genes that are positive predictors of immunotherapy response in human cancers. Our findings suggest that short in-frame deletions within BRAF exon 12 and MAP2K1 exons 2 and 3 in UDV595E cases are alternative MAPK-pathway activating events that may have significant therapeutic implications for selecting first-line treatment for canine UC. We developed a simple, cost-effective capillary electrophoresis genotyping assay for detection of these deletions in parallel with the BRAF V595E mutation. The identification of these deletion events in dogs offers a compelling cross-species platform in which to study the relationship between somatic alteration, protein conformation, and therapeutic sensitivity.

Transcriptomic profiling in canines and humans reveals cancer specific gene modules and biological mechanisms common to both species.

Understanding relationships between spontaneous cancer in companion (pet) canines and humans can facilitate biomarker and drug development in both species. Towards this end we developed an experimental-bioinformatic protocol that analyzes canine transcriptomics data in the context of existing human data to evaluate comparative relevance of canine to human cancer. We used this protocol to characterize five canine cancers: melanoma, osteosarcoma, pulmonary carcinoma, B- and T-cell lymphoma, in 60 dogs. We applied an unsupervised, iterative clustering method that yielded five co-expression modules and found that each cancer exhibited a unique module expression profile. We constructed cancer models based on the co-expression modules and used the models to successfully classify the canine data. These canine-derived models also successfully classified human tumors representing the same cancers, indicating shared cancer biology between canines and humans. Annotation of the module genes identified cancer specific pathways relevant to cells-of-origin and tumor biology. For example, annotations associated with melanin production (PMEL, GPNMB, and BACE2), synthesis of bone material (COL5A2, COL6A3, and COL12A1), synthesis of pulmonary surfactant (CTSH, LPCAT1, and NAPSA), ribosomal proteins (RPL8, RPS7, and RPLP0), and epigenetic regulation (EDEM1, PTK2B, and JAK1) were unique to melanoma, osteosarcoma, pulmonary carcinoma, B- and T-cell lymphoma, respectively. In total, 152 biomarker candidates were selected from highly expressing modules for each cancer type. Many of these biomarker candidates are under-explored as drug discovery targets and warrant further study. The demonstrated transferability of classification models from canines to humans enforces the idea that tumor biology, biomarker targets, and associated therapeutics, discovered in canines, may translate to human medicine.

Comparative Assessment of Pesticide Exposures in Domestic Dogs and Their Owners Using Silicone Passive Samplers and Biomonitoring.

Pesticides are used extensively in residential settings for lawn maintenance and in homes to control household pests including application directly on pets to deter fleas and ticks. Pesticides are commonly detected in the home environment where people and pets can be subject to chronic exposure. Due to increased interest in using companion animals as sentinels for human environmental health studies, we conducted a comparative pesticide exposure assessment in 30 people and their pet dogs to determine how well silicone wristbands and silicone dog tags can predict urinary pesticide biomarkers of exposure. Using targeted gas chromatography-mass spectrometry analyses, we quantified eight pesticides in silicone samplers and used a suspect screening approach for additional pesticides. Urine samples were analyzed for 15 pesticide metabolite biomarkers. Several pesticides were detected in >70% of silicone samplers including permethrin, N,N-diethyl-meta-toluamide (DEET), and chlorpyrifos. Significant and positive correlations were observed between silicone sampler levels of permethrin and DEET with their corresponding urinary metabolites (rs = 0.50-0.96, p < 0.05) in both species. Significantly higher levels of fipronil were observed in silicone samplers from participants who reported using flea and tick products containing fipronil on their dog. This study suggests that people and their dogs have similar pesticide exposures in a home environment.

Transcriptome annotation reveals minimal immunogenetic diversity among Wyoming toads, Anaxyrus baxteri.

Briefly considered extinct in the wild, the future of the Wyoming toad (Anaxyrus baxteri) continues to rely on captive breeding to supplement the wild population. Given its small natural geographic range and history of rapid population decline at least partly due to fungal disease, investigation of the diversity of key receptor families involved in the host immune response represents an important conservation need. Population decline may have reduced immunogenetic diversity sufficiently to increase the vulnerability of the species to infectious diseases. Here we use comparative transcriptomics to examine the diversity of toll-like receptors and major histocompatibility complex (MHC) sequences across three individual Wyoming toads. We find reduced diversity at MHC genes compared to bufonid species with a similar history of bottleneck events. Our data provide a foundation for future studies that seek to evaluate the genetic diversity of Wyoming toads, identify biomarkers for infectious disease outcomes, and guide breeding strategies to increase genomic variability and wild release successes.

Somatic inactivating PTPRJ mutations and dysregulated pathways identified in canine malignant melanoma by integrated comparative genomic analysis.

Canine malignant melanoma, a significant cause of mortality in domestic dogs, is a powerful comparative model for human melanoma, but little is known about its genetic etiology. We mapped the genomic landscape of canine melanoma through multi-platform analysis of 37 tumors (31 mucosal, 3 acral, 2 cutaneous, and 1 uveal) and 17 matching constitutional samples including long- and short-insert whole genome sequencing, RNA sequencing, array comparative genomic hybridization, single nucleotide polymorphism array, and targeted Sanger sequencing analyses. We identified novel predominantly truncating mutations in the putative tumor suppressor gene PTPRJ in 19% of cases. No BRAF mutations were detected, but activating RAS mutations (24% of cases) occurred in conserved hotspots in all cutaneous and acral and 13% of mucosal subtypes. MDM2 amplifications (24%) and TP53 mutations (19%) were mutually exclusive. Additional low-frequency recurrent alterations were observed amidst low point mutation rates, an absence of ultraviolet light mutational signatures, and an abundance of copy number and structural alterations. Mutations that modulate cell proliferation and cell cycle control were common and highlight therapeutic axes such as MEK and MDM2 inhibition. This mutational landscape resembles that seen in BRAF wild-type and sun-shielded human melanoma subtypes. Overall, these data inform biological comparisons between canine and human melanoma while suggesting actionable targets in both species.

A comprehensive genomic history of extinct and living elephants.

Elephantids are the world's most iconic megafaunal family, yet there is no comprehensive genomic assessment of their relationships. We report a total of 14 genomes, including 2 from the American mastodon, which is an extinct elephantid relative, and 12 spanning all three extant and three extinct elephantid species including an ∼120,000-y-old straight-tusked elephant, a Columbian mammoth, and woolly mammoths. Earlier genetic studies modeled elephantid evolution via simple bifurcating trees, but here we show that interspecies hybridization has been a recurrent feature of elephantid evolution. We found that the genetic makeup of the straight-tusked elephant, previously placed as a sister group to African forest elephants based on lower coverage data, in fact comprises three major components. Most of the straight-tusked elephant's ancestry derives from a lineage related to the ancestor of African elephants while its remaining ancestry consists of a large contribution from a lineage related to forest elephants and another related to mammoths. Columbian and woolly mammoths also showed evidence of interbreeding, likely following a latitudinal cline across North America. While hybridization events have shaped elephantid history in profound ways, isolation also appears to have played an important role. Our data reveal nearly complete isolation between the ancestors of the African forest and savanna elephants for ∼500,000 y, providing compelling justification for the conservation of forest and savanna elephants as separate species.

PTPN11 mutations in canine and human disseminated histiocytic sarcoma.

In humans, histiocytic sarcoma (HS) is an aggressive cancer involving histiocytes. Its rarity and heterogeneity explain that treatment remains a challenge. Sharing high clinical and histopathological similarities with human HS, the canine HS is conversely frequent in specific breeds and thus constitutes a unique spontaneous model for human HS to decipher the genetic bases and to explore therapeutic options. We identified sequence alterations in the MAPK pathway in at least 63.9% (71/111) of HS cases with mutually exclusive BRAF (0.9%; 1/111), KRAS (7.2%; 8/111) and PTPN11 (56.75%; 63/111) mutations concentrated at hotspots common to human cancers. Recurrent PTPN11 mutations are associated to visceral disseminated HS subtype in dogs, the most aggressive clinical presentation. We then identified PTPN11 mutations in 3/19 (15.7%) human HS patients. Thus, we propose PTPN11 mutations as key events for a specific subset of human and canine HS: the visceral disseminated form. Finally, by testing drugs targeting the MAPK pathway in eight canine HS cell lines, we identified a better anti-proliferation activity of MEK inhibitors than PTPN11 inhibitors in canine HS neoplastic cells. In combination, these results illustrate the relevance of naturally affected dogs in deciphering genetic mechanisms and selecting efficient targeted therapies for such rare and aggressive cancers in humans.

Comparative Exposure Assessment Using Silicone Passive Samplers Indicates That Domestic Dogs Are Sentinels To Support Human Health Research.

Silicone wristbands are promising passive samplers to support epidemiological studies in characterizing exposure to organic contaminants; however, investigating associated health risks remains challenging because of the latency period for many chronic diseases that take years to manifest. Dogs provide valuable insights as sentinels for exposure-related human disease because they share similar exposures in the home, have shorter life spans, share many clinical/biological features, and have closely related genomes. Here, we evaluated exposures among pet dogs and their owners using silicone dog tags and wristbands to determine if contaminant levels were correlated with validated exposure biomarkers. Significant correlations between measures on dog tags and wristbands were observed (rs = 0.38-0.90; p < 0.05). Correlations with their respective urinary biomarkers were stronger in dog tags compared to that in human wristbands (rs = 0.50-0.71; p < 0.01) for several organophosphate esters. This supports the value of using silicone bands with dogs to investigate health impacts on humans from shared exposures.

Genome-wide DNA copy number analysis and targeted transcriptional analysis of canine histiocytic malignancies identifies diagnostic signatures and highlights disruption of spindle assembly complex.

Canine histiocytic malignancies (HM) are rare across the general dog population, but overrepresented in certain breeds, such as Bernese mountain dog and flat-coated retriever. Accurate diagnosis relies on immunohistochemical staining to rule out histologically similar cancers with different prognoses and treatment strategies (e.g., lymphoma and hemangiosarcoma). HM are generally treatment refractory with overall survival of less than 6 months. A lack of understanding regarding the mechanisms of disease development and progression hinders development of novel therapeutics. While the study of human tumors can benefit veterinary medicine, the rarity of the suggested orthologous disease (dendritic cell sarcoma) precludes this. This study aims to improve the understanding of underlying disease mechanisms using genome-wide DNA copy number and gene expression analysis of spontaneous HM across several dog breeds. Extensive DNA copy number disruption was evident, with losses of segments of chromosomes 16 and 31 detected in 93% and 72% of tumors, respectively. Droplet digital PCR (ddPCR) evaluation of these regions in numerous cancer specimens effectively discriminated HM from other common round cell tumors, including lymphoma and hemangiosarcoma, resulting in a novel, rapid diagnostic aid for veterinary medicine. Transcriptional analysis demonstrated disruption of the spindle assembly complex, which is linked to genomic instability and reduced therapeutic impact in humans. A key signature detected was up-regulation of Matrix Metalloproteinase 9 (MMP9), supported by an immunohistochemistry-based assessment of MMP9 protein levels. Since MMP9 has been linked with rapid metastasis and tumor aggression in humans, the data in this study offer a possible mechanism of aggression in HM.

Exome sequencing of lymphomas from three dog breeds reveals somatic mutation patterns reflecting genetic background.

Lymphoma is the most common hematological malignancy in developed countries. Outcome is strongly determined by molecular subtype, reflecting a need for new and improved treatment options. Dogs spontaneously develop lymphoma, and the predisposition of certain breeds indicates genetic risk factors. Using the dog breed structure, we selected three lymphoma predisposed breeds developing primarily T-cell (boxer), primarily B-cell (cocker spaniel), and with equal distribution of B- and T-cell lymphoma (golden retriever), respectively. We investigated the somatic mutations in B- and T-cell lymphomas from these breeds by exome sequencing of tumor and normal pairs. Strong similarities were evident between B-cell lymphomas from golden retrievers and cocker spaniels, with recurrent mutations in TRAF3-MAP3K14 (28% of all cases), FBXW7 (25%), and POT1 (17%). The FBXW7 mutations recurrently occur in a specific codon; the corresponding codon is recurrently mutated in human cancer. In contrast, T-cell lymphomas from the predisposed breeds, boxers and golden retrievers, show little overlap in their mutation pattern, sharing only one of their 15 most recurrently mutated genes. Boxers, which develop aggressive T-cell lymphomas, are typically mutated in the PTEN-mTOR pathway. T-cell lymphomas in golden retrievers are often less aggressive, and their tumors typically showed mutations in genes involved in cellular metabolism. We identify genes with known involvement in human lymphoma and leukemia, genes implicated in other human cancers, as well as novel genes that could allow new therapeutic options.

Genomic profiling of canine mast cell tumors identifies DNA copy number aberrations associated with KIT mutations and high histological grade.

Mast cell tumor (MCT) is the most common skin malignancy of domestic dogs and presents with a widely variable clinical behavior. Although activating KIT mutations are present in approximately 20% of canine MCTs, molecular etiology is largely unknown for the majority of this cancer. Characterization of genomic alterations in canine MCTs may identify genomic regions and/or genes responsible for their development and progression, facilitating the discovery of new therapeutic targets and improved clinical management of this heterogeneous cancer. We performed genome-wide DNA copy number analysis of 109 primary MCTs derived from three popular canine breeds (the Boxer, Labrador Retriever, and Pug) as well as nontarget breeds using oligonucleotide array comparative genomic hybridization (oaCGH). We demonstrated a stepwise accumulation of numerical DNA copy number aberrations (CNAs) as tumor grade increases. DNA sequencing analysis revealed that KIT mutations were found less frequently in the Pug tumors and were strongly associated with high histological grade. Tumors with KIT mutations showed genome-wide aberrant copy number profiles, with frequent CNAs involving genes in the p53 and RB pathways, whereas CNAs were very limited in tumors with wild-type KIT. We evaluated the presence of four CNAs to predict aggressive tumor phenotypes. This approach predicted aggressive tumors with a sensitivity of 78-94% and specificity of 88-93%, when using oaCGH and droplet digital PCR platforms. Further investigation of genome regions identified in this study may lead to the development of a molecular tool for classification and prognosis, as well as identification of therapeutic target molecules.

Genome-wide association study identifies shared risk loci common to two malignancies in golden retrievers.

Dogs, with their breed-determined limited genetic background, are great models of human disease including cancer. Canine B-cell lymphoma and hemangiosarcoma are both malignancies of the hematologic system that are clinically and histologically similar to human B-cell non-Hodgkin lymphoma and angiosarcoma, respectively. Golden retrievers in the US show significantly elevated lifetime risk for both B-cell lymphoma (6%) and hemangiosarcoma (20%). We conducted genome-wide association studies for hemangiosarcoma and B-cell lymphoma, identifying two shared predisposing loci. The two associated loci are located on chromosome 5, and together contribute ~20% of the risk of developing these cancers. Genome-wide p-values for the top SNP of each locus are 4.6×10-7 and 2.7×10-6, respectively. Whole genome resequencing of nine cases and controls followed by genotyping and detailed analysis identified three shared and one B-cell lymphoma specific risk haplotypes within the two loci, but no coding changes were associated with the risk haplotypes. Gene expression analysis of B-cell lymphoma tumors revealed that carrying the risk haplotypes at the first locus is associated with down-regulation of several nearby genes including the proximal gene TRPC6, a transient receptor Ca2+-channel involved in T-cell activation, among other functions. The shared risk haplotype in the second locus overlaps the vesicle transport and release gene STX8. Carrying the shared risk haplotype is associated with gene expression changes of 100 genes enriched for pathways involved in immune cell activation. Thus, the predisposing germ-line mutations in B-cell lymphoma and hemangiosarcoma appear to be regulatory, and affect pathways involved in T-cell mediated immune response in the tumor. This suggests that the interaction between the immune system and malignant cells plays a common role in the tumorigenesis of these relatively different cancers.

The genome of the green anole lizard and a comparative analysis with birds and mammals.

The evolution of the amniotic egg was one of the great evolutionary innovations in the history of life, freeing vertebrates from an obligatory connection to water and thus permitting the conquest of terrestrial environments. Among amniotes, genome sequences are available for mammals and birds, but not for non-avian reptiles. Here we report the genome sequence of the North American green anole lizard, Anolis carolinensis. We find that A. carolinensis microchromosomes are highly syntenic with chicken microchromosomes, yet do not exhibit the high GC and low repeat content that are characteristic of avian microchromosomes. Also, A. carolinensis mobile elements are very young and diverse-more so than in any other sequenced amniote genome. The GC content of this lizard genome is also unusual in its homogeneity, unlike the regionally variable GC content found in mammals and birds. We describe and assign sequence to the previously unknown A. carolinensis X chromosome. Comparative gene analysis shows that amniote egg proteins have evolved significantly more rapidly than other proteins. An anole phylogeny resolves basal branches to illuminate the history of their repeated adaptive radiations.

Comparative cytogenetic characterization of primary canine melanocytic lesions using array CGH and fluorescence in situ hybridization.

Melanocytic lesions originating from the oral mucosa or cutaneous epithelium are common in the general dog population, with up to 100,000 diagnoses each year in the USA. Oral melanoma is the most frequent canine neoplasm of the oral cavity, exhibiting a highly aggressive course. Cutaneous melanocytomas occur frequently, but rarely develop into a malignant form. Despite the differential prognosis, it has been assumed that subtypes of melanocytic lesions represent the same disease. To address the relative paucity of information about their genomic status, molecular cytogenetic analysis was performed on the three recognized subtypes of canine melanocytic lesions. Using array comparative genomic hybridization (aCGH) analysis, highly aberrant distinct copy number status across the tumor genome for both of the malignant melanoma subtypes was revealed. The most frequent aberrations included gain of dog chromosome (CFA) 13 and 17 and loss of CFA 22. Melanocytomas possessed fewer genome wide aberrations, yet showed a recurrent gain of CFA 20q15.3-17. A distinctive copy number profile, evident only in oral melanomas, displayed a sigmoidal pattern of copy number loss followed immediately by a gain, around CFA 30q14. Moreover, when assessed by fluorescence in situ hybridization (FISH), copy number aberrations of targeted genes, such as gain of c-MYC (80 % of cases) and loss of CDKN2A (68 % of cases), were observed. This study suggests that in concordance with what is known for human melanomas, canine melanomas of the oral mucosa and cutaneous epithelium are discrete and initiated by different molecular pathways.

Genome-wide assessment of recurrent genomic imbalances in canine leukemia identifies evolutionarily conserved regions for subtype differentiation.

Leukemia in dogs is a heterogeneous disease with survival ranging from days to years, depending on the subtype. Strides have been made in both human and canine leukemia to improve classification and understanding of pathogenesis through immunophenotyping, yet classification and choosing appropriate therapy remains challenging. In this study, we assessed 123 cases of canine leukemia (28 ALLs, 24 AMLs, 25 B-CLLs, and 46 T-CLLs) using high-resolution oligonucleotide array comparative genomic hybridization (oaCGH) to detect DNA copy number alterations (CNAs). For the first time, such data were used to identify recurrent CNAs and inclusive genes that may be potential drivers of subtype-specific pathogenesis. We performed predictive modeling to identify CNAs that could reliably differentiate acute subtypes (ALL vs. AML) and chronic subtypes (B-CLL vs. T-CLL) and used this model to differentiate cases with up to 83.3 and 95.8 % precision, respectively, based on CNAs at only one to three genomic regions. In addition, CGH datasets for canine and human leukemia were compared to reveal evolutionarily conserved copy number changes between species, including the shared gain of HSA 21q in ALL and ∼25 Mb of shared gain of HSA 12 and loss of HSA 13q14 in CLL. These findings support the use of canine leukemia as a relevant in vivo model for human leukemia and justify the need to further explore the conserved genomic regions of interest for their clinical impact.

Definition of the cattle killer cell Ig-like receptor gene family: comparison with aurochs and human counterparts.

Under selection pressure from pathogens, variable NK cell receptors that recognize polymorphic MHC class I evolved convergently in different species of placental mammal. Unexpectedly, diversified killer cell Ig-like receptors (KIRs) are shared by simian primates, including humans, and cattle, but not by other species. Whereas much is known of human KIR genetics and genomics, knowledge of cattle KIR is limited to nine cDNA sequences. To facilitate comparison of the cattle and human KIR gene families, we determined the genomic location, structure, and sequence of two cattle KIR haplotypes and defined KIR sequences of aurochs, the extinct wild ancestor of domestic cattle. Larger than its human counterpart, the cattle KIR locus evolved through successive duplications of a block containing ancestral KIR3DL and KIR3DX genes that existed before placental mammals. Comparison of two cattle KIR haplotypes and aurochs KIR show the KIR are polymorphic and the gene organization and content appear conserved. Of 18 genes, 8 are functional and 10 were inactivated by point mutation. Selective inactivation of KIR3DL and activating receptor genes leaves a functional cohort of one inhibitory KIR3DL, one activating KIR3DX, and six inhibitory KIR3DX. Functional KIR diversity evolved from KIR3DX in cattle and from KIR3DL in simian primates. Although independently evolved, cattle and human KIR gene families share important function-related properties, indicating that cattle KIR are NK cell receptors for cattle MHC class I. Combinations of KIR and MHC class I are the major genetic factors associated with human disease and merit investigation in cattle.

Comprehensive genomic characterization of five canine lymphoid tumor cell lines.

BACKGROUND: Leukemia/lymphoma cell lines have been critical in the investigation of the pathogenesis and therapy of hematological malignancies. While human LL cell lines have generally been found to recapitulate the primary tumors from which they were derived, appropriate characterization including cytogenetic and transcriptional assessment is crucial for assessing their clinical predictive value. RESULTS: In the following study, five canine LL cell lines, CLBL-1, Ema, TL-1 (Nody-1), UL-1, and 3132, were characterized using extensive immunophenotyping, karyotypic analysis, oligonucleotide array comparative genomic hybridization (oaCGH), and gene expression profiling. Genome-wide DNA copy number data from the cell lines were also directly compared with 299 primary canine round cell tumors to determine whether the cell lines represent primary tumors, and, if so, what subtype each most closely resembled. CONCLUSIONS: Based on integrated analyses, CLBL-1 was classified as B-cell lymphoma, Ema and TL-1 as T-cell lymphoma, and UL-1 as T-cell acute lymphoblastic leukemia. 3132, originally classified as a B-cell lymphoma, was reclassified as a histiocytic sarcoma based on characteristic cytogenomic properties. In combination, these data begin to elucidate the clinical predictive value of these cell lines which will enhance the appropriate selection of in vitro models for future studies of canine hematological malignancies.

Identification of three molecular and functional subtypes in canine hemangiosarcoma through gene expression profiling and progenitor cell characterization.

Canine hemangiosarcomas have been ascribed to an endothelial origin based on histologic appearance; however, recent findings suggest that these tumors may arise instead from hematopoietic progenitor cells. To clarify this ontogenetic dilemma, we used genome-wide expression profiling of primary hemangiosarcomas and identified three distinct tumor subtypes associated with angiogenesis (group 1), inflammation (group 2), and adipogenesis (group 3). Based on these findings, we hypothesized that a common progenitor may differentiate into the three tumor subtypes observed in our gene profiling experiment. To investigate this possibility, we cultured hemangiosarcoma cell lines under normal and sphere-forming culture conditions to enrich for tumor cell progenitors. Cells from sphere-forming cultures displayed a robust self-renewal capacity and exhibited genotypic, phenotypic, and functional properties consistent with each of the three molecular subtypes seen in primary tumors, including expression of endothelial progenitor cell (CD133 and CD34) and endothelial cell (CD105, CD146, and αvß3 integrin) markers, expression of early hematopoietic (CD133, CD117, and CD34) and myeloid (CD115 and CD14) differentiation markers in parallel with increased phagocytic capacity, and acquisition of adipogenic potential. Collectively, these results suggest that canine hemangiosarcomas arise from multipotent progenitors that differentiate into distinct subtypes. Improved understanding of the mechanisms that determine the molecular and phenotypic differentiation of tumor cells in vivo could change paradigms regarding the origin and progression of endothelial sarcomas.

Genomic profiling reveals extensive heterogeneity in somatic DNA copy number aberrations of canine hemangiosarcoma.

Canine hemangiosarcoma is a highly aggressive vascular neoplasm associated with extensive clinical and anatomical heterogeneity and a grave prognosis. Comprehensive molecular characterization of hemangiosarcoma may identify novel therapeutic targets and advanced clinical management strategies, but there are no published reports of tumor-associated genome instability and disrupted gene dosage in this cancer. We performed genome-wide microarray-based somatic DNA copy number profiling of 75 primary intra-abdominal hemangiosarcomas from five popular dog breeds that are highly predisposed to this disease. The cohort exhibited limited global genomic instability, compared to other canine sarcomas studied to date, and DNA copy number aberrations (CNAs) were predominantly of low amplitude. Recurrent imbalances of several key cancer-associated genes were evident; however, the global penetrance of any single CNA was low and no distinct hallmark aberrations were evident. Copy number gains of dog chromosomes 13, 24, and 31, and loss of chromosome 16, were the most recurrent CNAs involving large chromosome regions, but their relative distribution within and between cases suggests they most likely represent passenger aberrations. CNAs involving CDKN2A, VEGFA, and the SKI oncogene were identified as potential driver aberrations of hemangiosarcoma development, highlighting potential targets for therapeutic modulation. CNA profiles were broadly conserved between the five breeds, although subregional variation was evident, including a near twofold lower incidence of VEGFA gain in Golden Retrievers versus other breeds (22 versus 40 %). These observations support prior transcriptional studies suggesting that the clinical heterogeneity of this cancer may reflect the existence of multiple, molecularly distinct subtypes of canine hemangiosarcoma.

Canine prostate cancer cell line (Probasco) produces osteoblastic metastases in vivo.

BACKGROUND: In 2012, over 240,000 men were diagnosed with prostate cancer and over 28,000 died from the disease. Animal models of prostate cancer are vital to understanding its pathogenesis and developing therapeutics. Canine models in particular are useful due to their similarities to late-stage, castration-resistant human disease with osteoblastic bone metastases. This study established and characterized a novel canine prostate cancer cell line that will contribute to the understanding of prostate cancer pathogenesis. METHODS: A novel cell line (Probasco) was derived from a mixed breed dog that had spontaneous prostate cancer. Cell proliferation and motility were analyzed in vitro. Tumor growth in vivo was studied by subcutaneous, intratibial, and intracardiac injection of Probasco cells into nude mice. Tumors were evaluated by bioluminescent imaging, Faxitron radiography, µCT, and histology. RT-PCR and genome-wide DNA copy number profiling were used to characterize the cell line. RESULTS: The Probasco cells grew in vitro (over 75 passages) and were tumorigenic in nude mice. Probasco cells expressed high levels of BMP2, CDH1, MYOF, FOLH1, RUNX2, and SMAD5 modest CXCL12, SLUG, and BMP, and no PTHrP mRNA. Following intracardiac injection, Probasco cells metastasized primarily to the appendicular skeleton, and both intratibial and intracardiac injections produced osteoblastic tumors in bone. Comparative genomic hybridization demonstrated numerous DNA copy number aberrations throughout the genome, including large losses and gains in multiple chromosomes. CONCLUSIONS: The Probasco prostate cancer cell line will be a valuable model to investigate the mechanisms of prostate cancer pathogenesis and osteoblastic bone metastases.