Assessment of Canine Mast Cell Tumor Mortality Risk Based on Clinical, Histologic, Immunohistochemical, and Molecular Features.

Mast cell tumor (MCT) is a frequent cutaneous neoplasm in dogs that is heterogeneous in clinical presentation and biological behavior, with a variable potential for recurrence and metastasis. Accurate prediction of clinical outcomes has been challenging. The study objective was to develop a system for classification of canine MCT according to the mortality risk based on individual assessment of clinical, histologic, immunohistochemical, and molecular features. The study included 149 dogs with a histologic diagnosis of cutaneous or subcutaneous MCT. By univariate analysis, MCT metastasis and related death was significantly associated with clinical stage ( P < .0001, rP = -0.610), history of tumor recurrence ( P < .0001, rP = -0.550), Patnaik ( P < .0001, rP = -0.380) and Kiupel grades ( P < .0001, rP = -0.500), predominant organization of neoplastic cells ( P < .0001, rP = -0.452), mitotic count ( P < .0001, rP = -0.325), Ki-67 labeling index ( P < .0001, rP = -0.414), KITr pattern ( P = .02, rP = 0.207), and c-KIT mutational status ( P < .0001, rP = -0.356). By multivariate analysis with Cox proportional hazard model, only 2 features were independent predictors of overall survival: an amendment of the World Health Organization clinical staging system (hazard ratio [95% CI]: 1.824 [1.210-4.481]; P = .01) and a history of tumor recurrence (hazard ratio [95% CI]: 9.250 [2.158-23.268]; P < .001]. From these results, we propose an amendment of the WHO staging system, a method of risk analysis, and a suggested approach to clinical and laboratory evaluation of dogs with cutaneous MCT.

Checkpoint Inhibitors in Dogs: Are We There Yet?

Immune checkpoint inhibitors (ICI) have revolutionised cancer treatment in people. Immune checkpoints are important regulators of the body's reaction to immunological stimuli. The most studied immune checkpoint molecules are programmed death (PD-1) with its ligand (PD-L1) and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) with its ligands CD80 (B7-1) and CD86 (B7-2). Certain tumours can evade immunosurveillance by activating these immunological checkpoint targets. These proteins are often upregulated in cancer cells and tumour-infiltrating lymphocytes, allowing cancer cells to evade immune surveillance and promote tumour growth. By blocking inhibitory checkpoints, ICI can help restore the immune system to effectively fight cancer. Several studies have investigated the expression of these and other immune checkpoints in human cancers and have shown their potential as therapeutic targets. In recent years, there has been growing interest in studying the expression of immune checkpoints in dogs with cancer, and a few small clinical trials with ICI have already been performed on these species. Emerging studies in veterinary oncology are centred around developing and validating canine-targeted antibodies. Among ICIs, anti-PD-1 and anti-PD-L1 treatments stand out as the most promising, mirroring the success in human medicine over the past decade. Nevertheless, the efficacy of caninized antibodies remains suboptimal, especially for canine oral melanoma. To enhance the utilisation of ICIs, the identification of predictive biomarkers for treatment response and the thorough screening of individual tumours are crucial. Such endeavours hold promise for advancing personalised medicine within veterinary practice, thereby improving treatment outcomes. This article aims to review the current research literature about the expression of immune checkpoints in canine cancer and the current results of ICI treatment in dogs.

Case report: Suspected plasmablastic lymphoma in a dog resembling the clinical presentation in humans.

Plasmablastic lymphoma (PBL) is a rare form of lymphoma in people. PBL originates from plasmablasts and usually presents with swelling/mass in the mouth/neck. A 7-year-old Mongrel dog was presented for a large oral and neck mass. Cytology and histopathology were suggestive of a round cell tumor that was suspected to be lymphoma. An immunohistochemical (IHC) stain panel showed positive for CD18, thus supporting the diagnosis of round cell tumor, but negative for T- and B-cell lymphomas, CD3, CD20, and PAX-5. Other markers including cytokeratin AE1/3 (for epithelial cell origin), CD31 (for endothelial cells), SOX10 (for melanoma), IBa-1 (for histiocytic sarcoma), and CD117 (for mast cell tumor) were all negative. MUM-1 (for plasma cell differentiation) was strongly positive and CD79a (B cell and plasma cells) was also scantly positive. Based on the histopathology and immunohistochemistry results in combination with the clinical presentation, a suspected diagnosis of PBL was made. As per available literature, this is perhaps the first highly suspected case of PBL in a dog.

Repurposing Drugs in Small Animal Oncology.

Repurposing drugs in oncology consists of using off-label drugs that are licensed for various non-oncological medical conditions to treat cancer. Repurposing drugs has the advantage of using drugs that are already commercialized, with known mechanisms of action, proven safety profiles, and known toxicology, pharmacokinetics and pharmacodynamics, and posology. These drugs are usually cheaper than new anti-cancer drugs and thus more affordable, even in low-income countries. The interest in repurposed anti-cancer drugs has led to numerous in vivo and in vitro studies, with some promising results. Some randomized clinical trials have also been performed in humans, with certain drugs showing some degree of clinical efficacy, but the true clinical benefit for most of these drugs remains unknown. Repurposing drugs in veterinary oncology is a very new concept and only a few studies have been published so far. In this review, we summarize both the benefits and challenges of using repurposed anti-cancer drugs; we report and discuss the most relevant studies that have been previously published in small animal oncology, and we suggest potential drugs that could be clinically investigated for anti-cancer treatment in dogs and cats.

Laboratory diagnosis of canine uroperitoneum based on cellular and biochemical characteristics of serum and abdominal fluid.

BACKGROUND: Literature on the laboratory diagnosis of uroperitoneum is scarce, and it is mostly based on the biochemical findings of cavitary fluid and serum. Cell count and protein concentrations measurements are rarely used and available studies on this subject are based on a relatively small cohort of individuals. OBJECTIVES: We aimed to use a large sample pool of dogs to establish cutoff points for biochemical analytes in cavitary fluids and serum for the diagnosis of uroperitoneum. We also sought to evaluate the general classification of these cavitary fluids. METHODS: In a retrospective and prospective study, 180 canine abdominal effusion cases were evaluated, 30 of which were uroperitoneum (uroperitoneum group, UG) and 150 with other etiologies (non-uroperitoneum group, NUG). RESULTS: The results showed that 83.3% of UG and 12.7% of NUG abdominal fluid cases were not classified as transudates or exudates. The use of specific cutoffs for fluid creatinine concentrations (≥2.1 mg/dL) and fluid:serum creatinine ratios (Cf: Cs ≥ 1.25) in these unclassified effusions resulted in an accuracy of 99.0% for the laboratory diagnosis of uroperitoneum. CONCLUSIONS: The adoption of a new set of criteria and cutoffs based on the combination of parameters such as TP, TNCC, fluid creatinine and Cf: Cs improves the diagnosis of uroperitoneum in dogs.

Inclusion of fibroblasts and collagen fibrils in the cytologic grading of canine cutaneous mast cell tumors.

BACKGROUND: Fibroblasts and/or collagen fibrils have not been included in previous cytologic grading schemes of canine mast cell tumors (MCTs), and their association with biological behavior is broadly debated. OBJECTIVES: This study aimed to evaluate the cytologic findings of canine MCT, with emphasis on the microenvironment, and propose a novel cytologic grading system correlated with mortality and histologic grade. MATERIAL AND METHODS: Cytology smears of canine cutaneous MCTs were retrospectively reviewed and compared with their histopathologic counterparts using Cohen´s Kappa test. One-year survival rates were also compared with the cytologic and histopathologic variables using Pearson´s correlation test. RESULTS: From 92 first-occurrence canine cutaneous MCTs, the five features most associated with mortality were selected for a new grading system. The five features were cytoplasmic granulation, fibroblast and/or collagen fibril concentrations, and the presence of mitotic figures, multinucleation, and karyomegaly. Among concordant histopathologic and cytologic cases (ie, the same grades using both systems), mortality rates were 2.6% (1/38) for low-grade and 71.4% (10/14) for high-grade cases (P < 0.001, chi-square). For false-negative and false-positive results, mortality rates were 33% (1/3) and 45% (5/11), respectively (P = 0.707). CONCLUSIONS: Unlike the Camus cytologic grading system, the present amendment excluded binucleation and included fibroblasts and/ or collagen fibrils, which in higher concentrations were associated with increased survival and a low histopathologic grade. Cytologic grading with the inclusion of fibroblast and collagen fibril concentrations correlated with survival, as did the Camus cytologic and Kiupel histopathologic grades; however, further studies are needed to confirm the prognostic value of this novel cytologic grading scheme.

Immunolabelling of Acetylated Histones 3 and 4 and Histone Deacetylases 1, 2 and 6 in Canine Soft Tissue Sarcomas.

The objective of this study was to investigate the immunolabelling of acetylated histones and histone desacetylase (HDAC) enzymes in canine soft tissue sarcomas (STSs) and to correlate them with histological and clinical features in order to identify possible prognostic and therapeutic targets in these neoplasms. Fifteen canine STS samples were evaluated and were submitted to immunohistochemistry for acetylated histones 3 (H3) and 4 (H4) and deacetylating enzymes (HDAC1, HDAC2 and HDAC6). Intense immunolabelling of H4 was seen in comparison with H3. A strong positive correlation was observed between the H3 intensity score and the number of mitotic figures (P = 0.004, r = 0.7). Intense immunolabelling of HDAC1 was found in comparison to the expression of HDAC2 and HDAC6 in the evaluated STSs. This finding suggests that HDAC1 may be a potential target for HDAC inhibitors in STSs in dogs.

Epidemiological study of canine transmissible venereal tumor (CTVT) in Brazil, 2000-2020.

Canine transmissible venereal tumor (CTVT) is a contagious neoplasm, mainly transmitted through coitus. This round cell mesenchymal tumor is common in Brazil, often located in the genitalia although extragenital presentations may also occur, such as cutaneous, oral, and nasal forms. The objective of this study was to perform an epidemiological analysis of CTVT from published data in the recent academic literature to systematically demonstrate the distribution of CTVT in Brazil, identify the frequency of this neoplasm and its main diagnostic tests, and characterize its main clinical manifestations in Brazil. For such purpose, it was analyzed the scientific publications with cases of CTVT in Brazil, in English or Portuguese, published between 2000-2020. The CTVT was identified in 19 Brazilian states plus the Federal District, totaling 3,622 cases across the national territory, with the largest number of cases recorded in the Southeast region. The cytological exam was the most used for the diagnosis of CTVT (89.2 %), followed by histopathological (37.8 %) and immunohistochemistry (13.5 %)1 . Predominant epidemiological aspects of CTVT identified in the study were: Mixed breed dogs (75.2 %), females (62.5 %), in adulthood (between 2 and 7 years) and dogs with free extra outdoor access (91.1 %). Genital presentation was the most frequent in the literature (86 %), followed by cutaneous (21.8 %), nasal (10 %), oral and lymph nodes presentations (10-5 %) and less frequent manifestations as ocular and anal/perianal (< 5 %). CTVT is a neoplasm widely distributed in Brazil, highly frequent and with several forms of clinical presentation, which can be underdiagnosed if there is no adequate knowledge of this tumor and its epidemiological characteristics. The extragenital manifestations of the neoplasm need further studies for its better characterization and more precise definition of its frequencies.

LOPH: a novel chemotherapeutic protocol for feline high-grade multicentric or mediastinal lymphoma, developed in an area endemic for feline leukemia virus.

OBJECTIVES: The aim of this study was to establish the safety and efficacy of a novel multidrug lomustine-based chemotherapeutic protocol for cats with high-grade multicentric or mediastinal lymphoma, in an area endemic for feline leukemia virus (FeLV). METHODS: This prospective study included owned cats, diagnosed (cytologically) with multicentric or mediastinal lymphoma and treated with the LOPH (lomustine, vincristine [Oncovin; Antibióticos do Brasil], prednisolone and hydroxydaunorubicin [doxorubicin]) protocol. A complete blood count was performed before every chemotherapy session and any significant abnormalities recorded as possible related toxicities. Median survival time (MST) and disease-free interval were estimated by Kaplan-Meier curves. RESULTS: Twenty-one cats were included in this study. Nineteen (90.5%) tested positive for FeLV and were therefore considered to have persistent viremia. Complete response was reported in 81% (n = 17/21), while three had partial remission and one had no response. Seven cats finished the induction protocol within 20-31 weeks (23.1 ± 4.5; median 20) and all seven received a maintenance protocol. The MST (lymphoma-related survival) for the 21 cats was 214 days. The MST was 214 days for cats with mediastinal lymphoma (n = 13), but it was not reached for multicentric lymphoma (n = 8; P = 0.9). The MST of cats with persistent FeLV antigenemia was 171 days. Grade I anorexia and vomiting occurred in 19% of the cats (n = 4/21). Hematologic toxicity was found in 100% of the cats at some point during their treatment, but it was mostly grade I or II. Neutropenia, thrombocytopenia and anemia occurred in 16/21, 21/21 and 15/21 cats, respectively. CONCLUSIONS AND RELEVANCE: The LOPH protocol was well tolerated by cats with lymphoma and persistent FeLV viremia, and resulted in a better MST than similar studies with other protocols. Novel studies and controlled trials are necessary in order to evaluate the efficacy of different protocols according to the lymphoma subtype, anatomic form and FeLV status.

Recurrent horizontal transfer identifies mitochondrial positive selection in a transmissible cancer.

Autonomous replication and segregation of mitochondrial DNA (mtDNA) creates the potential for evolutionary conflict driven by emergence of haplotypes under positive selection for 'selfish' traits, such as replicative advantage. However, few cases of this phenomenon arising within natural populations have been described. Here, we survey the frequency of mtDNA horizontal transfer within the canine transmissible venereal tumour (CTVT), a contagious cancer clone that occasionally acquires mtDNA from its hosts. Remarkably, one canine mtDNA haplotype, A1d1a, has repeatedly and recently colonised CTVT cells, recurrently replacing incumbent CTVT haplotypes. An A1d1a control region polymorphism predicted to influence transcription is fixed in the products of an A1d1a recombination event and occurs somatically on other CTVT mtDNA backgrounds. We present a model whereby 'selfish' positive selection acting on a regulatory variant drives repeated fixation of A1d1a within CTVT cells.

Somatic evolution and global expansion of an ancient transmissible cancer lineage.

The canine transmissible venereal tumor (CTVT) is a cancer lineage that arose several millennia ago and survives by "metastasizing" between hosts through cell transfer. The somatic mutations in this cancer record its phylogeography and evolutionary history. We constructed a time-resolved phylogeny from 546 CTVT exomes and describe the lineage's worldwide expansion. Examining variation in mutational exposure, we identify a highly context-specific mutational process that operated early in the cancer's evolution but subsequently vanished, correlate ultraviolet-light mutagenesis with tumor latitude, and describe tumors with heritable hyperactivity of an endogenous mutational process. CTVT displays little evidence of ongoing positive selection, and negative selection is detectable only in essential genes. We illustrate how long-lived clonal organisms capture changing mutagenic environments, and reveal that neutral genetic drift is the dominant feature of long-term cancer evolution.

Surgical stress and postoperative complications related to regional and radical mastectomy in dogs.

BACKGROUND: Surgery is the treatment of choice for regional control of mammary neoplasms in female dogs. Various surgical techniques may be used, as long as mammary gland anatomy, lymphatic drainage, and known prognostic factors are respected. The purpose of this study was to compare surgical stress-including duration of surgery, nociception and hematological changes-and postoperative complications in dogs undergoing regional and unilateral radical mastectomy. Eighteen dogs were selected for each technique. Postoperative pain (nociception), hematological changes, and postoperative complications were compared between the two groups. RESULTS: The group treated with radical mastectomy had a longer surgical duration, showed more intense physiological changes, achieved higher scores on nociception scales, and experienced more postoperative complications. CONCLUSION: Compared to regional mastectomy, radical mastectomy was associated with longer surgical duration, greater nociceptive stimulus, greater surgical stress, and higher incidence of postoperative complications in dogs. Although evaluation of long-term results was not a goal of this study, it is suggested that postoperative recovery and patient quality of life should be considered when choosing a surgical approach for treating mammary tumors in dogs.