Targeting canine mammary neoplastic epithelial cells with a reengineered anthrax toxin: first study.

Mammary tumors are the most frequent type of neoplasms in intact female dogs. New therapies that target neoplastic cells without affecting normal cells are highly sought. The Bacillus anthracis toxin has been reengineered to target tumor cells that express urokinase plasminogen activators and metalloproteinases. In previous studies carried out in our laboratory, the reengineered anthrax toxin had inhibitory effects on canine oral mucosal melanoma and canine osteosarcoma cells. In this study, five canine neoplastic epithelial cell lines (four adenocarcinomas and one adenoma) and one non-neoplastic canine mammary epithelial cell line were treated with different concentrations of reengineered anthrax toxin components. Cell viability was quantified using an MTT assay and half-maximal inhibitory concentration (IC50) values. Cell lines were considered sensitive when the IC50 was lower than 5000 ng/ml. One canine mammary adenocarcinoma cell line and one mammary adenoma cell line showed significantly decreased viability after treatment, whereas the non-neoplastic cell line was resistant. We conclude that the reengineered anthrax toxin may be considered a targeted therapy for canine mammary neoplasms while preserving normal canine mammary epithelial cells.

High lysyl oxidase expression is an indicator of poor prognosis in dogs with cutaneous mast cell tumours.

Mast cell tumour (MCT) is one of the most frequent skin tumours in dogs. Due to their unpredictable biological behaviour, MCTs often cause several therapeutic frustrations, leading to investigation regarding prognostic markers. Lysyl oxidase (LOX) is an enzyme that promotes extracellular matrix stability and contributes to cell migration, angiogenesis and epithelial-mesenchymal transition. Its expression positively correlates with poor prognoses in several human and canine mammary cancers. The aim of this study was to characterise the immunohistochemical expression of LOX in MCT samples and compare it with histological grading and post-surgical survival. Twenty-six tumours were submitted to immunohistochemistry for LOX expression evaluation. All samples were positive for LOX, with variable percentages of cytoplasmic and nuclear positivity. Cytoplasmic positivity was significantly higher in high-grade MCTs (P = .0297). Our results indicate that high expression of cytoplasmic LOX in neoplastic mast cells is an indicator of poor prognosis for canine cutaneous MCTs.

Diagnosis, Prognosis, and Treatment of Canine Hemangiosarcoma: A Review Based on a Consensus Organized by the Brazilian Association of Veterinary Oncology, ABROVET.

Hemangiosarcoma is a mesenchymal neoplasm originating in the endothelial cells of blood vessels; they can be classified as non-visceral and visceral types. Non-visceral hemangiosarcomas can affect the skin, subcutaneous tissues, and muscle tissues; visceral hemangiosarcomas can affect the spleen, liver, heart, lungs, kidneys, oral cavity, bones, bladder, uterus, tongue, and retroperitoneum. Among domestic species, dogs are most affected by cutaneous HSA. Cutaneous HSA represents approximately 14% of all HSA diagnosed in this species and less than 5% of dermal tumors, according to North American studies. However, Brazilian epidemiological data demonstrate a higher prevalence, which may represent 27 to 80% of all canine HSAs and 13.9% of all skin neoplasms diagnosed in this species. Cutaneous HSA most commonly affects middle-aged to elderly dogs (between 8 and 15 years old), with no gender predisposition for either the actinic or non-actinic forms. The higher prevalence of cutaneous HSA in some canine breeds is related to lower protection from solar radiation, as low skin pigmentation and hair coverage lead to greater sun exposure. Actinic changes, such as solar dermatosis, are frequent in these patients, confirming the influence of solar radiation on the development of this neoplasm. There are multiple clinical manifestations of hemangiosarcoma in canines. The diagnostic approach and staging classification of cutaneous HSAs are similar between the different subtypes. The definitive diagnosis is obtained through histopathological analysis of incisional or excisional biopsies. Cytology can be used as a presurgical screening test; however, it has little diagnostic utility in cases of HSA because there is a high risk of blood contamination and sample hemodilution. Surgery is generally the treatment of choice for dogs with localized non-visceral HSA without evidence of metastatic disease. Recently, electrochemotherapy (ECT) has emerged as an alternative therapy for the local ablative treatment of different neoplastic types; the use of radiotherapy for the treatment of dogs with cutaneous HSA is uncommon. There is greater consensus in the literature regarding the indications for adjuvant chemotherapy in subcutaneous and muscular HSA; doxorubicin is the most frequently used antineoplastic agent for subcutaneous and muscular subtypes and can be administered alone or in combination with other drugs. Other therapies include antiangiogenic therapy, photodynamic therapy, the association of chemotherapy with the metronomic dose, targeted therapies, and natural products. The benefits of these therapies are presented and discussed. In general, the prognosis of splenic and cardiac HSA is unfavorable. As a challenging neoplasm, studies of new protocols and treatment modalities are necessary to control this aggressive disease.

Diagnosis, Prognosis and Treatment of Canine Cutaneous and Subcutaneous Mast Cell Tumors.

Mast cell tumors (MCTs) are hematopoietic neoplasms composed of mast cells. It is highly common in dogs and is extremely important in the veterinary oncology field. It represents the third most common tumor subtype, and is the most common malignant skin tumor in dogs, corresponding to 11% of skin cancer cases. The objective of this critical review was to present the report of the 2nd Consensus meeting on the Diagnosis, Prognosis, and Treatment of Canine Cutaneous and Subcutaneous Mast Cell Tumors, which was organized by the Brazilian Association of Veterinary Oncology (ABROVET) in August 2021. The most recent information on cutaneous and subcutaneous mast cell tumors in dogs is presented and discussed.

Intercellular interactions between mast cells and stromal fibroblasts obtained from canine cutaneous mast cell tumours.

Mast cell tumours (MCTs) are the most frequent malignant skin neoplasm in dogs. Due to the difficulty in purifying large numbers of canine neoplastic mast cells, relatively little is known about their properties. A reproducible in vitro model is needed to increase the understanding about the phenotype and functional properties of neoplastic mast cells. In the present study, we describe the establishment of primary cocultures of neoplastic mast cells from canine cutaneous MCTs and cancer-associated fibroblasts. We confirmed the inability of canine neoplastic mast cells to remain viable for long periods in vitro without the addition of growth factors or in vivo passages in mice. Using a transwell system, we observed that mast cell viability was significantly higher when there is cell-to-cell contact in comparison to non-physical contact conditions and that mast cell viability was significantly higher in high-grade than in low-grade derived primary cultures. Moreover, the use of conditioned medium from co-cultured cells led to a significantly higher tumoral mast cell viability when in monoculture. Signalling mechanisms involved in these interactions might be attractive therapeutic targets to block canine MCT progression and deserve more in-depth investigations.

Galectin-3 immunolabelling correlates with BCL2 expression in canine cutaneous mast cell tumours.

Mast cell tumour (MCT) is the most frequent skin neoplasm in dogs. These tumours are characterised by variable behaviour and clinical presentation that make prognosis an important and challenging task in the veterinary practice. Galectin-3 (Gal-3) is known to influence several biological processes that are important in the cancer context and has been described as a prognostic marker for several human cancers. The aim of the present work was to characterise Gal-3 immunolabelling in canine cutaneous MCTs and to investigate its value as a prognostic marker for the disease. Thirty-four random cases of canine cutaneous MCT that were surgically treated with wide margins were included in this study. Gal-3 expression was evaluated using immunohistochemistry and the results were compared with the expression of apoptosis-related proteins, Ki67 index, histopathological grades, mortality due to the disease and post-surgical survival. The majority of the MCTs (65.8%) were positive for Gal-3. Gal-3 immunolabelling was variable among the samples (2.7%-86.8% of the neoplastic cells). The protein was located in the cytoplasm or in the cytoplasm and the nucleus. Gal-3 positivity was correlated with BCL2 expression (P < 0.001; r = 0.604), but not with Ki67 and BAX. No significant differences were detected between histological grades or in the survival analysis. Gal-3 expression correlates with BCL2 expression in MCTs. Although an efficient marker for several human neoplasms, the results presented herein suggest that Gal-3 immunolabelling is not an independent prognostic indicator for this disease.

Inhibitory Effects of a Reengineered Anthrax Toxin on Canine and Human Osteosarcoma Cells.

Canine and human osteosarcomas (OSA) share similarities. Novel therapies are necessary for these tumours. The Bacillus anthracis toxin was reengineered to target and kill cells with high expressions of matrix metalloproteinases (MMPs) and urokinase plasminogen activator (uPA). Since canine OSA express MMPs and uPA, we assessed whether the reengineered toxin could show efficacy against these tumours. Two OSA cell lines (canine D17 and human MG63) and a non-neoplastic canine osteoblastic cell line (COBS) were used. Cells were treated with different concentrations of the reengineered anthrax toxin and cell viability was quantified using MTT assay. The cell cycle, apoptosis, and necrosis were analysed by flow cytometry. The wound-healing assay was performed to quantify the migration capacity of treated cells. D17 and MG63 cells had significantly decreased viability after 24 h of treatment. Cell cycle analysis revealed that OSA cells underwent apoptosis when treated with the toxin, whereas COBS cells arrested in the G1 phase. The wound-healing assay showed that D17 and MG63 cells had a significantly reduced migration capacity after treatment. These results point for the first time towards the in vitro inhibitory effects of the reengineered anthrax toxin on OSA cells; this reengineered toxin could be further tested as a new therapy for OSA.

Inhibitory Effects of a Reengineered Anthrax Toxin on Canine Oral Mucosal Melanomas.

Canine oral mucosal melanomas (OMM) are the most common oral malignancy in dogs and few treatments are available. Thus, new treatment modalities are needed for this disease. Bacillus anthracis (anthrax) toxin has been reengineered to target tumor cells that express urokinase plasminogen activator (uPA) and metalloproteinases (MMP-2), and has shown antineoplastic effects both, in vitro and in vivo. This study aimed to evaluate the effects of a reengineered anthrax toxin on canine OMM. Five dogs bearing OMM without lung metastasis were included in the clinical study. Tumor tissue was analyzed by immunohistochemistry for expression of uPA, uPA receptor, MMP-2, MT1-MMP and TIMP-2. Animals received either three or six intratumoral injections of the reengineered anthrax toxin prior to surgical tumor excision. OMM samples from the five dogs were positive for all antibodies. After intratumoral treatment, all dogs showed stable disease according to the canine Response Evaluation Criteria in Solid Tumors (cRECIST), and tumors had decreased bleeding. Histopathology has shown necrosis of tumor cells and blood vessel walls after treatment. No significant systemic side effects were noted. In conclusion, the reengineered anthrax toxin exerted inhibitory effects when administered intratumorally, and systemic administration of this toxin is a promising therapy for canine OMM.

Adrenalectomy in dogs: retrospective study of 13 cases performed in Universidade Anhembi Morumbi Veterinary Hospital (2012-2015) / Adrenalectomia em cães: estudo retrospectivo de 13 casos realizados no hospital veterinário da Universidade Anhembi Morumbi (2012 a 2015)

Adrenalectomy is the most appropriate treatment for unilateral adrenal tumors. This study aimed at describing the epidemiological characteristics and perioperative behavior of canine patients submitted to adrenalectomy at Anhembi Morumbi Veterinary Hospital. Out of 13 dogs, eight were pure breeds and five were mixed breeds; 12 females, aged 9.5 ± 2.5 years old. Regarding the tumors, seven were located on the right and histopathological analysis revealed cortical adenoma in 11 and adenocarcinoma in only two dogs. Two cases had hypercortisolism recurrence associated with hyperplasia in the contralateral adrenal, as confirmed by ACTH stimulation test. The results of this study indicate that adrenalectomy is a safe procedure with few perioperative complications, despite the possibility of hypercortisolism recurrence.(AU)

Adrenalectomia é o tratamento mais indicado para neoplasias adrenais unilaterais. Objetivou-se descrever os aspectos epidemiológicos e o comportamento perioperatório em cães submetidos a adrenalectomia no Hospital Veterinário Anhembi Morumbi. Dos 13 casos, oito eram cães de raças puras e cinco eram SRD; 12 eram fêmeas e de idade de 9,5 ± 2,5 anos. Das massas adrenais, sete eram em lado direito. Os exames histopatológicos revelaram adenoma cortical em 11 cães e adenocarcinoma em dois. Dois casos recidivaram a condição de hipercortisolemia, associada à hiperplasia, na adrenal contralateral, confirmados pelo teste de estimulação por ACTH. Concluiu-se que a adrenalectomia é um procedimento seguro, com poucas complicações perioperatórias, ressalvando-se a possiblidade de recidiva do quadro de hipercortisolismo.(AU)

Comparative Aspects of Canine Melanoma.

Melanomas are malignant neoplasms originating from melanocytes. They occur in most animal species, but the dog is considered the best animal model for the disease. Melanomas in dogs are most frequently found in the buccal cavity, but the skin, eyes, and digits are other common locations for these neoplasms. The aim of this review is to report etiological, epidemiological, pathological, and molecular aspects of melanomas in dogs. Furthermore, the particular biological behaviors of these tumors in the different body locations are shown. Insights into the therapeutic approaches are described. Surgery, chemotherapy, radiotherapy, immunotherapy, and the outcomes after these treatments are presented. New therapeutic perspectives are also depicted. All efforts are geared toward better characterization and control of malignant melanomas in dogs, for the benefit of these companion animals, and also in an attempt to benefit the treatment of human melanomas.

Environmental risk factors related to the development of canine non-Hodgkin's lymphoma / Fatores de risco ambientais relacionados ao desenvolvimento do linfoma não Hodgkin canino

This research aimed to investigate the possible risk factors associated with the development of canine non-Hodgkin's lymphoma. Owners of 83 dogs with non-Hodgkin's lymphoma and of 84 healthy dogs answered an epidemiological questionnaire. Dogs who lived outside of the house and within 100 meters of busy streets or avenues (defined as more than 50 vehicles per minute) had a higher risk for developing the disease (OR: 3.1, 95% CI: 1.4-6.9, P=0.005). These results suggest that air pollution derived from vehicle traffic may be associated with the development of canine non-Hodgkin's lymphoma.

Este trabalho teve como objetivo investigar os possíveis fatores de risco ambientais, associados com o desenvolvimento de linfoma não-Hodgkin nos cães. Um questionário epidemiológico foi aplicado aos proprietários de 83 cães com linfoma não-Hodgkin e 84 proprietários de cães saudáveis. Os cães que viviam permanentemente no lado de fora da casa e em torno de 100 metros de ruas movimentadas ou avenidas (mais de 50 veículos por minuto) tiveram um maior risco de desenvolvimento da doença (OR: 3,1, IC 95%: 1,4-6,9, P=0,005). Esses resultados sugerem que a poluição do ar oriunda do tráfego veicular pode estar associada com o desenvolvimento de linfomas não-Hodgkin canino.