Background: The immune system plays an important role in the multifactorial biologic system during the development of neoplasias. However, the involvement of the inflammatory response in the promotion/control of malignant cells is still controversial, and the cell subsets and the mechanisms involved are poorly investigated. The goal of this study was to characterize the clinical-pathological status and the immunophenotyping profile of tumor infiltrating lymphocytes and their association with the animal survival rates in canine mammary carcinomas.Methods: Fifty-one animals with mammary carcinomas, classified as carcinomas in mixed tumors-MC-BMT = 31 and carcinomas-MC = 20 were submitted to systematic clinical-pathological analysis (tumor size; presence of lymph node and pulmonary metastasis; clinical stage; histological grade; inflammatory distribution and intensity as well as the lymphocytic infiltrate intensity) and survival rates. Twenty-four animals (MC-BMT = 16 and MC = 8) were elected to the immunophenotypic study performed by flow cytometry.Results: Data analysis demonstrated that clinical stage II-IV and histological grade was I more frequent in MC-BMT as compared to MC. Univariate analysis demonstrated that the intensity of inflammation (moderate/intense) and the proportion of CD4⁺(≥ 66.7%) or CD8⁺T-cells (<33.3%) were not associated with worse survival rate. Multivariate analysis demonstrated that only lymphocytic infiltrate intensity ≥ 600 (P= 0.02) remained as independent prognostic factor. Despite the clinical manifestation, the lymphocytes represented the predominant cell type in the tumor infiltrate. The percentage of T-cells was higher in animals with MC-BMT without metastasis, while the percentage of B-lymphocytes was greater in animals with metastasized MC-BMT (P< 0.05). The relative percentage of CD4⁺T-cells was significantly greater in metastasized tumors (both MC-BMT and MC), (P< 0.05) while the proportion of CD8⁺T-cells was higher in MC-BMT without metastasis. Consequently, the CD4⁺/CD8⁺ratio was significantly increased in both groups with metastasis. Regardless of the tumor type, the animals with high proportions of CD4⁺and low CD8⁺T-cells had decreased survival rates.Conclusion: The intensity of lymphocytic infiltrate and probably the relative abundance of the CD4⁺and CD8⁺T-lymphocytes may represent important survival prognostic biomarkers for canine mammary carcinomas.

Mammary tumors are among the most common neoplastic processes in female dogs. Prostaglandin E2, the catalytic product of Cox-2, may promote tumor development and angiogenesis. It has been investigated in several human cancers and also correlated with the evolution of the disease. However, the clinical implications of tumor pathology require more investigation in veterinary medicine. Angiogenesis is essential for the growth and metastasis of major solid tumors and has been correlated with prognosis in human and canine breast cancer. The aim of this study was to evaluate Cox-2 expression and microvessel density in canine mammary carcinomas and to correlate them with overall survival of the animal. Cox-2 and angiogenesis were assessed by immunohistochemistry in 46 mammary carcinomas (19 ductal and 27 metaplastic) and in healthy mammary glands. To assess tumor angiogenesis, microvessel density (MVD) was determined by CD31 staining. Immunostaining revealed that 46/46 (100%) of the tumors were positive for Cox-2 and CD31, and there was no statistical difference among tumor types. Cox-2 protein expression correlated positively with CD31 staining ( r = 0.3742, P = .0104) but did not correlate significantly with tumor type. Longer overall survival was observed in metaplastic carcinomas ( P = .028), in tumors with low microvessel density ( P = .0002) and with low Cox-2 score ( P = .01). Our results demonstrate that increased microvessel density and increased Cox-2 expression were linearly related in the canine mammary tumors studied and were also related to worse prognosis and shorter overall survival. This suggests that Cox-2 inhibitors could be an alternative for the treatment and control of advanced neoplastic mammary disease in female dogs.

The objective of the present investigation was to study the expression of c-erbB-2 and MIB-1 and try to associate them with morphological features of the cell such as nuclear pleomorphism, mitotic count and histological grade in a series of 70 canine mammary gland tumors, 22 of them benign and 48 malignant. Tumors were collected at the Veterinary Hospital of UFMG (Brazil) and the Veterinary Faculty of Porto University (Portugal). c-erbB-2 expression was determined according to the guidelines provided by the manufacturer of the HercepTest system and nuclear pleomorphism, mitotic count and histological grade according the Elston and Ellis grading system. The HercepTest is the FDA-approved in vitro diagnostic test marketed by Dako. It is a semi-quantitative immunohistochemical assay used to determine overexpression of HER2 protein (human epidermal growth factor receptor) in breast cancer tissue. MIB-1 expression was also evaluated in 28 malignant tumors. Seventeen (35.4%) of the malignant tumors were positive for c-erbB-2 expression, which was positively associated with nuclear pleomorphism (P < 0.0001), histological grade (P = 0.0017) and mitotic count (P < 0.05). Nuclear pleomorphism also showed a positive association with MIB-1 index (P < 0.0001). These results suggest that some of the biological and morphological characteristics of the tumor are associated in canine mammary gland tumors, as also reported for human breast cancer. It was also possible to show that the immunoexpression of c-erbB-2 can be a factor in mammary carcinogenesis. This fact opens the possibility of using anti-c-erbB-2 antibodies in the treatment of canine mammary tumors.

Canine mammary neoplasms (CMNs) are the most frequent lesions and in female dogs. However, studies correlating pathological criteria with clinical evolution in female dogs with mammary neoplasms are scarce. The present study aims to present epidemiological, clinical-pathological and overall survival data to help establish the prognosis and understand the biological behavior of CMNs. A total of 1539 cases were included (85% malignant and 13% benign). Tumor size was an important prognostic factor and was associated with overall patient survival (P< 0.0001). Most dogs diagnosed with malignant neoplasms (83%) had initial clinical staging, although 17% had regional or distant metastases at the time of diagnosis and lower overall survival (P< 0.0001). Carcinoma in mixed tumor was the most frequent histological type and had a better prognosis. Solid carcinomas, micropapillary carcinomas and carcinosarcomas were considered histological types with aggressive biological behavior and were associated with a worse prognosis and lower overall survival (P< 0.0001).