Evaluating the Histologic Grade of Digital Squamous Cell Carcinomas in Dogs with Dark and Light Haircoat - A Comparative Study of the Invasive Front and Tumor Cell Budding Systems

Canine digital squamous cell carcinomas (CDSCC) are particularly aggressive when compared to their occurrence in other locations. Although these neoplasms are more frequently seen in dark-haired dogs, such as Giant Schnauzers, there are no data checking whether these tumors are histologically different between breeds. We histologically evaluated DSCC from 94 dogs. These were divided into two groups, namely, (1) dark-haired (N = 76) and (2) light-haired breeds (N = 18), further subdividing Group 1 into three subgroups, (1a) black breeds (n = 11), (1b) Schnauzers (n = 34) and (1c) black & tan breeds (n = 31). Adaptations from two different squamous cell carcinomas grading schemes from human and veterinary literature were used. Both systems showed significant differences when compared to Groups 1 and 2 in terms of final grade, invasive front keratinization, degree of invasion, nuclear pleomorphism, tumor cell budding, smallest tumor nest size and amount of tumor stroma. Group 2 was consistently better differentiated CDSCC than Group 1. However, there were no significant differences among the dark-haired breeds in any of the features evaluated. This study represents the first attempt to grade CDSCC while taking into account both phenotypical and presumptive genotypical haircoat color. In conclusion, CDSCC are not only more common in dark-haired dogs, they are also histologically more aggressive

First evidence of vertical Hepatozoon canis transmission in dogs in Europe

Background Hepatozoon canis is a protozoal agent that is known to be transmitted by oral uptake of H. canis-infected Rhipicephalus sanguineus sensu lato ticks in dogs. Vertical transmission of H. canis has only been described once in a study evaluating dogs from Japan. The aim of this study was to investigate the parasitological status of puppies from a bitch that had tested positive for Hepatozoon spp. prior to giving birth. Findings A 4-year-old, female, pregnant dog imported from Italy (Sardinia) to Germany showed clinical signs of lethargy and tachypnoea and tested positive for H. canis by PCR. The dog gave birth to eight puppies, one of which was stillborn and another that had to be reanimated. Haematology, buffy coat analysis and a biochemistry profile were performed for each dog. EDTA-blood of the surviving seven puppies and bone marrow, liver, spleen, amniotic fluid, and umbilical cord of the stillborn puppy was tested for the presence of Hepatozoon spp. by PCR. The mother and the seven surviving puppies tested positive for H. canis by PCR at day 62 post-partum. Gamonts were detected in all dogs by buffy coat evaluation. Haematological and biochemistry results revealed mild abnormalities. In the stillborn puppy, spleen, umbilical cord, and amniotic fluid were positive for H. canis. Conclusion The results confirm that vertical transmission is a possible route of H. canis infection in dogs, demonstrated by molecular detection of the pathogen in the stillborn puppy. In the seven surviving puppies, vertical transmission was the most likely transmission route. A potential impact of the level of parasitaemia on the health of puppies, as well as its pathogenesis, should be investigated further

Cross-species oncogenomics offers insight into human muscle-invasive bladder cancer

Background In humans, muscle-invasive bladder cancer (MIBC) is highly aggressive and associated with a poor prognosis. With a high mutation load and large number of altered genes, strategies to delineate key driver events are necessary. Dogs and cats develop urothelial carcinoma (UC) with histological and clinical similarities to human MIBC. Cattle that graze on bracken fern also develop UC, associated with exposure to the carcinogen ptaquiloside. These species may represent relevant animal models of spontaneous and carcinogen-induced UC that can provide insight into human MIBC. Results Whole-exome sequencing of domestic canine (n = 87) and feline (n = 23) UC, and comparative analysis with human MIBC reveals a lower mutation rate in animal cases and the absence of APOBEC mutational signatures. A convergence of driver genes (ARID1A, KDM6A, TP53, FAT1, and NRAS) is discovered, along with common focally amplified and deleted genes involved in regulation of the cell cycle and chromatin remodelling. We identify mismatch repair deficiency in a subset of canine and feline UCs with biallelic inactivation of MSH2. Bovine UC (n = 8) is distinctly different; we identify novel mutational signatures which are recapitulated in vitro in human urinary bladder UC cells treated with bracken fern extracts or purified ptaquiloside. Conclusion Canine and feline urinary bladder UC represent relevant models of MIBC in humans, and cross-species analysis can identify evolutionarily conserved driver genes. We characterize mutational signatures in bovine UC associated with bracken fern and ptaquiloside exposure, a human-linked cancer exposure. Our work demonstrates the relevance of cross-species comparative analysis in understanding both human and animal UC

Artificial Intelligence to Predict the BRAF V595E Mutation in Canine Urinary Bladder Urothelial Carcinomas.

In dogs, the BRAF mutation (V595E) is common in bladder and prostate cancer and represents a specific diagnostic marker. Recent advantages in artificial intelligence (AI) offer new opportunities in the field of tumour marker detection. While AI histology studies have been conducted in humans to detect BRAF mutation in cancer, comparable studies in animals are lacking. In this study, we used commercially available AI histology software to predict BRAF mutation in whole slide images (WSI) of bladder urothelial carcinomas (UC) stained with haematoxylin and eosin (HE), based on a training (n = 81) and a validation set (n = 96). Among 96 WSI, 57 showed identical PCR and AI-based BRAF predictions, resulting in a sensitivity of 58% and a specificity of 63%. The sensitivity increased substantially to 89% when excluding small or poor-quality tissue sections. Test reliability depended on tumour differentiation (p < 0.01), presence of inflammation (p < 0.01), slide quality (p < 0.02) and sample size (p < 0.02). Based on a small subset of cases with available adjacent non-neoplastic urothelium, AI was able to distinguish malignant from benign epithelium. This is the first study to demonstrate the use of AI histology to predict BRAF mutation status in canine UC. Despite certain limitations, the results highlight the potential of AI in predicting molecular alterations in routine tissue sections

Artificial Intelligence to Predict the BRAF V595E Mutation in Canine Urinary Bladder Urothelial Carcinomas

In dogs, the BRAF mutation (V595E) is common in bladder and prostate cancer and represents a specific diagnostic marker. Recent advantages in artificial intelligence (AI) offer new opportunities in the field of tumour marker detection. While AI histology studies have been conducted in humans to detect BRAF mutation in cancer, comparable studies in animals are lacking. In this study, we used commercially available AI histology software to predict BRAF mutation in whole slide images (WSI) of bladder urothelial carcinomas (UC) stained with haematoxylin and eosin (HE), based on a training (n = 81) and a validation set (n = 96). Among 96 WSI, 57 showed identical PCR and AI-based BRAF predictions, resulting in a sensitivity of 58% and a specificity of 63%. The sensitivity increased substantially to 89% when excluding small or poor-quality tissue sections. Test reliability depended on tumour differentiation (p < 0.01), presence of inflammation (p < 0.01), slide quality (p < 0.02) and sample size (p < 0.02). Based on a small subset of cases with available adjacent non-neoplastic urothelium, AI was able to distinguish malignant from benign epithelium. This is the first study to demonstrate the use of AI histology to predict BRAF mutation status in canine UC. Despite certain limitations, the results highlight the potential of AI in predicting molecular alterations in routine tissue sections

Cross-species oncogenomics offers insight into human muscle-invasive bladder cancer

Background In humans, muscle-invasive bladder cancer (MIBC) is highly aggressive and associated with a poor prognosis. With a high mutation load and large number of altered genes, strategies to delineate key driver events are necessary. Dogs and cats develop urothelial carcinoma (UC) with histological and clinical similarities to human MIBC. Cattle that graze on bracken fern also develop UC, associated with exposure to the carcinogen ptaquiloside. These species may represent relevant animal models of spontaneous and carcinogen-induced UC that can provide insight into human MIBC. Results Whole-exome sequencing of domestic canine (n = 87) and feline (n = 23) UC, and comparative analysis with human MIBC reveals a lower mutation rate in animal cases and the absence of APOBEC mutational signatures. A convergence of driver genes (ARID1A, KDM6A, TP53, FAT1, and NRAS) is discovered, along with common focally amplified and deleted genes involved in regulation of the cell cycle and chromatin remodelling. We identify mismatch repair deficiency in a subset of canine and feline UCs with biallelic inactivation of MSH2. Bovine UC (n = 8) is distinctly different; we identify novel mutational signatures which are recapitulated in vitro in human urinary bladder UC cells treated with bracken fern extracts or purified ptaquiloside. Conclusion Canine and feline urinary bladder UC represent relevant models of MIBC in humans, and cross-species analysis can identify evolutionarily conserved driver genes. We characterize mutational signatures in bovine UC associated with bracken fern and ptaquiloside exposure, a human-linked cancer exposure. Our work demonstrates the relevance of cross-species comparative analysis in understanding both human and animal UC

Evaluating the Histologic Grade of Digital Squamous Cell Carcinomas in Dogs with Dark and Light Haircoat—A Comparative Study of the Invasive Front and Tumor Cell Budding Systems

Canine digital squamous cell carcinomas (CDSCC) are particularly aggressive when compared to their occurrence in other locations. Although these neoplasms are more frequently seen in dark-haired dogs, such as Giant Schnauzers, there are no data checking whether these tumors are histologically different between breeds. We histologically evaluated DSCC from 94 dogs. These were divided into two groups, namely, (1) dark-haired (N = 76) and (2) light-haired breeds (N = 18), further subdividing Group 1 into three subgroups, (1a) black breeds (n = 11), (1b) Schnauzers (n = 34) and (1c) black &amp; tan breeds (n = 31). Adaptations from two different squamous cell carcinomas grading schemes from human and veterinary literature were used. Both systems showed significant differences when compared to Groups 1 and 2 in terms of final grade, invasive front keratinization, degree of invasion, nuclear pleomorphism, tumor cell budding, smallest tumor nest size and amount of tumor stroma. Group 2 was consistently better differentiated CDSCC than Group 1. However, there were no significant differences among the dark-haired breeds in any of the features evaluated. This study represents the first attempt to grade CDSCC while taking into account both phenotypical and presumptive genotypical haircoat color. In conclusion, CDSCC are not only more common in dark-haired dogs, they are also histologically more aggressive

Correlation of BRAF Variant V595E, Breed, Histological Grade and Cyclooxygenase-2 Expression in Canine Transitional Cell Carcinomas

The presence of BRAF variant V595E, as well as an increased cyclooxygenase-2 (COX-2) expression in canine transitional cell carcinoma (TCC) are well-described in the literature. The aim of the present study was to investigate the correlation between breed (terrier versus non-terrier dogs), histological grade, COX-2 expression, and BRAF mutation in canine TCC. Therefore, transmural TCC biopsies from 65 dogs (15 terriers, 50 non-terriers) were graded histologically into low- and high-grade. Immunohistochemical evaluation of the intensity of COX-2 expression was performed using an immunoreactive score (IRS). Exon 15 of chromosome 16 was examined for the BRAF variant c.1799T&gt;A by TaqMan&reg; SNP assay. TCC was low-grade in 20 cases (one terrier, 19 non-terriers) and high-grade in 45 cases (14 terriers, 31 non-terriers). Contrary to humans, histological grade was not significantly correlated to the intensity of COX-2 expression. BRAF mutation was detected in 11/15 (73%) TCC of terriers and in 18/50 (36%) TCC of non-terriers. Histological grade and BRAF mutation were not correlated significantly (p = 0.2912). Terriers had a considerably higher prevalence of high-grade tumors (p &lt; 0.0001), as well as of BRAF mutation (p &le; 0.05) compared to non-terriers. In non-terriers, neoplasms with BRAF mutation showed a significantly higher intensity of COX-2 expression than those without BRAF mutation (p &le; 0.05). In conclusion, in contrast to humans, testing for BRAF mutation in canine TCC is a sensitive diagnostic method especially in terriers (73%) and may be recommended as a screening test. However, evidence of BRAF mutation in canine TCC is not a predictor for the histological grade. Moreover, a positive correlation between histological grade and the intensity of COX-2 expression was not found. Further studies are necessary to clarify the clinical and prognostic relevance of the elevated intensity of COX-2 expression of TCC with BRAF mutation detected in non-terriers