Malignant Mammary Tumors Research Articles

Presence and Importance of Oxidative Stress Parameters in Malignant Mammary Gland Tumors in Dogs

Presence and Importance of Oxidative Stress Parameters in Malignant Mammary Gland Tumors in Dogs

Представления о болезни у пациенток со злокачественными и доброкачественными новообразованиями молочных желез

The paper reports variants of subjective illness perception in patients with benign and malignant mammary tumors. The study included 100 patients followed-up in the Vladimirsky Moscow Regional Research Cinical Institute (MONIKI) (50 patients with stage II–III breast cancer and 50 patients with fibrocystic breast changes). The study provided data on subjective illness concept variants for patients of both groups which defined the illness behavior of patients, including the non-compliant behavior. The results obtained may be used for development of disease prevention programs (primary and secondary prevention).

Leptin Modulates the Metastasis of Canine Inflammatory Mammary Adenocarcinoma Cells Through Downregulation of Lysosomal Protective Protein Cathepsin A (CTSA).

Canine malignant mammary gland tumors present with a poor prognosis due to metastasis to other organs, such as lung and lymph node metastases. Unlike in human studies where obesity has been shown to increase the risk of breast cancer, this has not been well studied in veterinary science. In our preliminary study, we discovered that leptin downregulated cathepsin A, which is responsible for lysosomal-associated membrane protein 2a (LAMP2a) degradation. LAMP2a is a rate-limiting factor in chaperone-mediated autophagy and is highly active in malignant cancers. Therefore, in this study, alterations in metastatic capacity through cathepsin A by leptin, which are secreted at high levels in the blood of obese patients, were investigated. We used a canine inflammatory mammary gland adenocarcinoma (CHMp) cell line cultured with RPMI-1640 and 10% fetal bovine serum. The samples were then subjected to real-time polymerase chain reaction, Western blot, immunocytochemistry, and lysosome isolation to investigate and visualize the metastasis and chaperone-mediated autophagy-related proteins. Results showed that leptin downregulated cathepsin A expression at both transcript and protein levels, whereas LAMP2a, the rate-limiting factor of chaperone-mediated autophagy, was upregulated by inhibition of LAMP2a degradation. Furthermore, leptin promoted LAMP2a multimerization through the lysosomal mTORC2 (mTOR complex 2)/PH domain and leucine rich repeat protein phosphatase 1 (PHLPP1)/AKT1 (Serine/threonine-protein kinase 1) pathway. These findings suggest that targeting leptin receptors can alleviate mammary gland cancer cell metastasis in dogs.

Prognostic utility of preoperative neutrophil–lymphocyte ratio in cats with malignant mammary tumors

Although studies have determined the neutrophil–lymphocyte ratio (NLR) to be a useful prognostic predictor in various human cancers, little is known regarding its preoperative prognostic significance in feline mammary tumors (FMTs). Thus, the present study aimed to determine the prognostic utility of preoperative peripheral blood NLR in malignant FMTs. Accordingly, medical records of 34 cats that had undergone mastectomy were searched, after which those histopathologically confirmed to have malignant FMTs were included. Univariate analysis was used to determine the effect of each variable on one-year survival time. Factors determined to be significant during univariate analysis were subsequently included for multivariate analysis. Receiver operating characteristic analysis was used to assess the accuracy of the tests for predicting one-year survival. Univariate analysis showed that a high NLR and large tumor size were significantly associated with shorter survival times (P < 0.01, for both). No significant differences in other factors were observed. Multivariate analysis showed that a high NLR (hazard ratio, 25.03; 95% confidence interval, 2.98–210.40; P < 0.01), but not tumor size and age, was an independent prognostic factor for poor outcomes. The sensitivity and specificity of the NLR (cut-off, 5.67) for predicting one-year survival were 93.8% and 88.9%, respectively. This study suggests that the NLR could potentially be used as a preoperative prognostic factor in FMTs. Preoperative NLR may help to establish therapeutic modalities by allowing prognostic evaluation among cats with mammary tumors.

Increased programmed death ligand (PD-L1) and cytotoxic T-lymphocyte antigen-4 (CTLA-4) expression is associated with metastasis and poor prognosis in malignant canine mammary gland tumours

Aberrant expression of immune check point molecules, programmed death ligand (PD-L1) and cytotoxic T-lymphocyte antigen-4 (CTLA-4) has been reported in many human cancers with increased protein and gene expression correlated with an aggressive behaviour in some neoplasms. Additionally, PD-L1 blockade has been shown to be an effective therapy for some human cancers. Canine mammary gland tumours have previously been shown to produce PD-L1 protein, but there are no previous studies investigating CTLA-4 in these common canine neoplasms. The present study investigated protein and gene expression of PD-L1 and CTLA-4 using immunohistochemistry and RT-PCR in 41 histologically-malignant, outcome-known CMGTs. The PD-L1 and CTLA-4 immunostaining scores of the mammary gland tumours that subsequently metastasised were significantly higher than those of tumours which did not metastasise (PD-L1: p = 0.005, CTLA-4: p = 0.003). Gene expression of PD-L1 and CTLA-4 was also significantly higher in tumours which subsequently metastasised (PD-L1: p = 0.023, CTLA-4: p = 0.022). Further, higher PD-L1 or CTLA-4 immunostaining scores correlated with shorter survival times of dogs (PD-L1: rs = - 0.42, p = 0.008, CTLA-4: rs = - 0.4, p = 0.01) while PD-L1 immunostaining was independently prognostic of survival time (Δ F = 4.9, p = 0.035). These findings suggest that higher protein and gene expression of PD-L1 and CTLA-4 by tumour cells increases the chances of metastasis and measuring these proteins may predict likely neoplasm behaviour. Additionally, if increased expression of these proteins promotes metastasis, blocking PD-L1 or CTLA-4 may be beneficial to treat canine mammary gland tumours.

Significant association of serum autoantibodies to TYMS, HAPLN1 and IGFBP5 with early stage canine malignant mammary tumours.

Canine mammary tumours (CMTs) are the most prevalent neoplasms in female dogs. Despite the high incidence of such tumours, a lack of easily accessible biomarkers still impedes early diagnosis of malignant CMTs. Herein we identify thymidylate synthetase (TYMS), hyaluronan and proteoglycan link protein 1 (HAPLN1) and insulin-like growth factor-binding protein 5 (IGFBP5) as CMT antigens eliciting corresponding autoantibodies in CMT cases. We establish enzyme-linked immunosorbent assays (ELISAs) to detect autoantibodies to TYMS (TYMS-AAb), HAPLN1 (HAPLN1-AAb) and IGFBP5 (IGFBP5-AAb) in sera from 81 dogs with malignant CMTs (41 in Stage I), 24 with benign CMTs and 35 healthy controls. Levels of all the three autoantibodies are elevated in the malignant group compared with the healthy or the benign group; notably, the elevated autoantibody levels significantly correlate with the stage-I CMTs. For discriminating malignant CMTs from healthy control, the area under curve (AUC) of TYMS-AAb is 0.694 with specificity of 82.9% and sensitivity of 50.6%. The AUC of utilising HAPLN1-AAb for distinguishing the stage-I CMTs from healthy controls is 0.711 with specificity of 77.1% and sensitivity of 58.5%. In differentiating malignant CMTs from the benign, the AUC of IGFBP5-AAb reaches 0.696 with specificity of 70.8% and sensitivity of 67.9%, and a combination of IGFBP5-AAb and TYMS-AAb increases the AUC to 0.72. Finally, the AUC of combined HAPLN1-AAb and IGFBP5-AAb in discriminating the stage-I CMTs from the benign achieves 0.731. Collectively, this study highlights a significant association of the three serum autoantibodies with early stage malignant CMTs.

Bioaccumulation of pesticide contaminants in tissue matrices of dogs suffering from malignant canine mammary tumors in Punjab, India

The unprecedented application of pesticides in Punjab, India during green revolution has lead to an environmental crisis due to the accumulation of persistent organic and pesticide pollutants in the environment and biota of this region. The present study aimed at estimating the abundance of pesticide contaminants in three biological matrices of 36 dogs suffering from malignant canine mammary tumor (mCMT) and 6 tumor free control dogs from Punjab, India. Presence of individual and total pesticides in canine biological samples, age and bodyweight of canine patients was assessed as a potential risk factor for mCMT using logistic regression analysis. Chi-square test was employed to determine tissue-specific accumulations of individual pesticides. Spearman's correlation coefficient was estimated to determine the association between the levels of total pesticides in different tissue matrices and with age and bodyweight of mCMT cases. Gas chromatography-ECD analysis of serum, mammary tissue and adjoining mammary adipose tissue revealed fourteen different pesticides including γ-HCH, α-HCH, dieldrin, aldrin, heptachlor, butachlor, p,p-DDT, o,p-DDT, p,p-DDD, p,p-DDE, L-cyhalothrin, permethrin, fipronil, and fenitrothion. Heptachlor, γ-HCH, aldrin and p,p-DDT were more frequently detected, whereas, p,p-DDE and o,p-DDT were the least common. Differential accumulation of pesticides in tissue matrices, particularly between serum and mammary tissue/adipose tissue was observed. We could not find any association between the total pesticide concentrations among serum, mammary tissue and mammary adipose tissue in mCMT cases. We found that the odds for individual pesticide for serum, mammary tissue and adipose tissue were associated with high uncertainties; however, the total pesticide concentration in mammary tissue was near non-significantly associated with higher risk of mCMT with low uncertainty. Statistically non-significant higher odds of CMT occurrence with increase in age was noticed No association between the concentration of total pesticides in different matrices and age and bodyweight of canine subjects was found.

Immunohistochemical study of IGF-I and IGF-II expression in canine mammary tumours: Prognostic and diagnostic role.

Mammary tumours are among the most common tumours in dogs and are of interest due to their similarities to human breast tumours. Insulin-like growth factors (IGFs) are considered important in cell growth and development. The aim of this study was to investigate the immunohistochemical expression of IGF-I and IGF-II in benign and malignant canine mammary tumours. In this study, 10 benign and 10 malignant mammary tumours from the archives of the Department of Pathology were used, and five normal breast tissues were used as controls. It was observed that the expression of IGF-I and IGF-II was low to absent in benign tumours and increased in malignant tumours. The expression of IGF-II was higher than that of IGF-I. This study showed that IGF-I and IGF-II can be used as criteria for malignancy in canine mammary tumours. The results also indicate that IGF-I and IGF-II may be used as early diagnostic markers, and their inhibition may be used for the treatment of canine and human mammary tumours in the future.

Canine Mammary Gland Cancer Stem Cell and its Potential Role in Malignant Biologic Behavior

BACKGROUND:Canine mammary gland cancers are the most prevalent malignancies in dogs. There are different challenges regarding management of these cancers in dogs and human, one hypothesis is related to small cellular subset of tumor mass called cancer stem cell. These cells are therapy resistant and cause metastasis and relapse even after primary successful treatment. The well-identified phenotypes for detecting this population are ALDH1+/CD44+/CD24-/Low biomarkers. OBJECTIVES: A study to evaluate existence of cancer stem cells in canine malignant mammary glands tumor and assess effects of these cells on clinicopathological parameters of tumors were designed. METHODS: In this study forty cases of canine mammary glands tumors were collected. All cases were tested via H&E and then Immunohistochemistry (IHC) methods. All samples were evaluated immunohis- tochemically for common markers of these tumor-initiating cells. Monoclonal antibodies against ALDH1, CD44 and CD24 were used. Some tumor aggressiveness-related parameters, including lymphovascular in- vasion, tumor grades and histotypes were assessed. RESULTS: The present study revealed that 17.5% of cases were enriched with cancer stem cells and all of them were diagnosed as grade II and III (P ≤ 0.05). Other findings showed all cancer stem cell-positive cases were significantly lymphovascular invasion positive (P ≤ 0.05). The most common histotypes in this research were tubular, tubulopapillary and intraductal carcinomas. CONCLUSIONS: Our results illustrated that cancer stem cells can be considered as reliable prognostic factors to predict severity of malignant behavior of canine malignant mammary glands cancer, which is comparable with human breast cancer.

Decorin concentrations in canine normal and neoplastic mammary gland tissues.

In the present study, the concentration of decorin in canine normal and neoplastic mammary gland tissues was examined to understand the potential role of decorin in development and progression of canine mammary tumours. The homogenates of 48 mammary gland tumours (10 benign and 38 malignant) and 10 samples of normal canine mammary gland tissue were used in the study. The presence and quantification of decorin was examined in the homogenates using Western blot and specific canine ELISA. Western blotting confirmed the presence of decorin both in the normal mammary gland tissues and in the mammary gland tumours. The concentration of decorin was significantly higher (p<.05) in the benign tumours and non-metastatic malignant tumours than in the normal mammary gland. The concentration of decorin was significantly lower (p<.05) in the malignant tumours with metastasis to regional lymph nodes compared with benign tumours and non-metastatic malignant tumours. No significant differences were found in the level of decorin between the benign and the non-metastatic malignant tumours. Both the histological type of malignant tumours and the histological grade did not significantly affect the concentration of decorin. These findings suggest that neoplastic transformation in the canine mammary gland leads to increase in the decorin protein synthesis. The reducing decorin concentration in canine malignant mammary tumours appears to facilitate the metastatic spread of these tumours.

HER-2 positivity rate in dogs with mammary carcinoma: a systematic review and meta-analysis

Human epidermal growth factor receptor 2 (HER-2) plays an essential role in cell growth and survival. HER-2 overexpression occurs in 20-30% of human breast tumors and has prognostic value as it is associated with disease progression. HER-2 overexpression is also associated with tumor progression and metastasis in malignant mammary tumors of the canine. However, in the literature, different positivity classifications/scoring were used in the evaluation of HER-2 status, and there is no consensus in terms of scoring of HER-2 expressionin canine mammary tumors. In this study, it was aimed to estimate the HER-2 positivity rate by evaluating the results of the study using different positivity classifications by meta-analysis. In this context, by using ”HER-2 canine mammary tumor” keywords, Pubmed and Web of Science electronic databases were scanned until February 2019, and a total of 97 related studies were found. However, 20 of these studies were used for the analysis. Two different meta-analyses were performed to evaluate the HER-2 positivity status with “2+ and 3+” and “3+” scores. As a result, HER-2 positivity rates were determined at 25.87% and 25.99% for the studies using “2+ / 3+” scores and “3+” respectively for HER-2 positivity. Therefore, this result suggests that the rate of HER-2 positivity is similar between humans and dogs.

Intraoperative near-infrared imaging can identify canine mammary tumors, a spontaneously occurring, large animal model of human breast cancer.

IntroductionCurrent methods of intraoperative margin assessment in breast conserving surgery are impractical, unreliable, or time consuming. We hypothesized that intraoperative near-infrared (NIR) imaging with an FDA-approved NIR optical contrast agent could identify canine mammary tumors, a spontaneous large animal model of human breast cancer, during surgery.MethodsDogs with mammary tumors underwent a standard of care lumpectomy or mastectomy with wide surgical margins 20 hours after indocyanine green administration (3 mg/kg IV). During surgery, NIR imaging was performed on tumors and wound margins in situ and tumors and lymph nodes ex vivo. Following resection, the wound bed was examined for residual fluorescence. Fluorescence intensity was determined by signal-to-background ratio (SBR). All tumors, areas of residual fluorescence, and lymph nodes underwent histopathologic analysis.ResultsThere were 41 mammary tumors in 16 female dogs. Twenty tumors were malignant and 21 were benign. Twenty-eight tumors were fluorescent (mean SBR 1.5±0.2). Sensitivity of fluorescence for all malignant tumors was 80% (16/20) and 93.3% (14/15) for malignant tumors > 2 cm. Specificity for malignancy was low (< 2cm = 55%; > 2cm = 30%). Tumors > 2 cm were more likely to be fluorescent (OR 6.05, 95% CI 1.50–24.44, P = 0.011) but not more likely to be malignant (OR 3.09, 95% CI 0.86–11.14, P = 0.085) than tumors ≤ 2 cm. Four out of seven inguinal lymph nodes excised in the mastectomy specimen fluoresced. All four drained malignant tumors; however only 2/4 contained metastatic disease.ConclusionSystemic ICG accumulates reliably in malignant canine mammary tumors > 2 cm. Although no tumor margins fluoresced, a wider margin of normal tissue is removed in canine mastectomy, making direct comparisons with breast conserving surgery difficult. Targeted NIR imaging agents are likely required to improve detection of smaller tumors and improve the specificity of NIR imaging for residual disease and metastatic lymph node detection.

Prognostic significance of immunohistochemical markers and histological classification in malignant canine mammary tumours.

Canine mammary carcinoma represents a model for the study of human breast cancer, although the prognostic value of various clinical, histological and immunohistochemical parameters has shown contradictory results. A prospective study, through a 4‐year follow‐up, was performed in 77 patients with mammary carcinoma to analyse the association between histological diagnosis, grade of malignancy, peritumoral and vascular invasion. We have also performed immunohistochemistry for the expression of oestrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and cyclooxygenase‐2 (COX‐2) that define human biomarkers of disease progression and treatment response. An association between histological diagnosis and clinical stage was observed with a high proportion of complex carcinoma classified as stage I. There was a higher proportion of ER+/PR+/HER2− tumours in stage I. In contrast, triple‐negative tumours (ER−/PR−/HER2−) were found mainly in advanced clinical stages and were associated with vascular and peritumoral invasion. The tumours included in group VII (carcinosarcoma/adenosquamous carcinoma/other special types of carcinoma) had a higher expression of COX‐2. The univariate analysis showed that those patients with complex carcinoma had the lowest incidence of metastases and the highest probability of survival. In contrast, a high proportion of patients with anaplastic/inflammatory carcinoma developed metastases and showed the lowest probability of survival. In addition, the estimated survival time was shorter for those patients with triple‐negative tumours and those with high COX‐2 expression. However, in the multivariate analysis, only the peritumoral invasion maintained its prognostic significance. In conclusion, in our study anaplastic/inflammatory carcinomas had the worst prognosis with a high proportion of triple‐negative tumours in this category.

Circulating microRNA as biomarkers of canine mammary carcinoma in dogs.

BackgroundDifferentiating benign from canine malignant mammary tumors requires invasive surgical biopsy. Circulating microRNAs (miRNA) may represent promising minimally invasive cancer biomarkers in people and animals.ObjectivesTo evaluate the serum mRNA profile between dogs with and without mammary carcinoma, and to determine if any of these markers have prognostic significance.AnimalsTen healthy client‐owned female dogs (5 intact, 5 spayed) and 10 dogs with histologically confirmed mammary carcinoma were included; 9 were client‐owned, whereas 1 was a research colony dog.MethodsRetrospective study. Serum miRNA was evaluated by RNA deep‐sequencing (RNAseq) and digital droplet PCR (dPCR).Expression of candidate biomarkers miR‐18a, miR‐19b, miR‐29b, miR‐34c, miR‐122, miR‐125a, and miR‐181a was compared with clinical characteristics, including grade, metastasis, and survival.Results452 unique serum miRNAs were detected by RNAseq. Sixty‐five individual miRNAs were differentially expressed (>±1.5‐fold) and statistically significant between groups. Serum miR‐19b (P = .003) and miR‐125a (P < .001) were significantly higher in the mammary carcinoma group by dPCR. Both had high accuracy based on receiver operator characteristic area under the curve (0.930 for miR‐125a; 0.880 for miR‐19b). Circulating miR‐18a by RNAseq was significantly higher in mammary carcinoma dogs with histologic evidence of lymphatic invasion (P = 0.03). There was no significant association with any miRNA and survival or inflammatory status.Conclusions and Clinical ImportanceCirculating miRNAs are differentially expressed in dogs with mammary carcinoma. Serum miR‐19b and miR‐18a represent candidate biomarkers for diagnosis and prognosis, respectively.

The Expression of Selected Factors Related to T Lymphocyte Activity in Canine Mammary Tumors.

Crosstalk between neoplastic and immune cells in the tumor microenvironment (TME) influences the progression of disease in human and canine cancer patients. Given that canine mammary tumors are a useful model to study breast cancer biology, we aimed to evaluate the expression of genes associated with T lymphocyte activity in benign, malignant, and metastatic canine mammary tumors. Interestingly, metastatic tumors exhibit increased expression of CXCR3, CCR2, IL-4, IL-12p40, and IL-17. In particular, we focused on IL-17, a key interleukin associated with the Th17 lymphocyte phenotype. Th17 cells have been shown to play a contradictory role in tumor immunity. Although IL-17 showed a high expression in the metastatic tumors, the expression of RORγt, a crucial transcription factor for Th17 differentiation was barely detected. We further investigated IL-17 expression using immunohistochemistry, through which we confirmed the increased expression of this interleukin in malignant and metastatic mammary tumors. Finally, we compared the plasma levels of IL-17 in healthy and malignant mammary tumor-bearing dogs using ELISA but found no differences between the groups. Our data indicate that the IL-17 in metastatic tumors may be produced by other cell types, but not by Th17 lymphocytes. Overall, our results broaden the available knowledge on the interactions in canine mammary tumors and provide insight into the development of new therapeutic strategies, with potential benefits for human immune oncology.

Differential stromal reprogramming in benign and malignant naturally occurring canine mammary tumours identifies disease-modulating stromal components

While cancer-associated stroma (CAS) in malignant tumours is well described, stromal changes in benign forms of naturally occurring tumours remain poorly characterized. Spontaneous canine mammary carcinomas (mCA) are viewed as excellent models of human mCA. We have recently reported highly conserved stromal reprogramming between canine and human mCA based on transcriptome analysis of laser-capture-microdissected FFPE specimen. To identify stromal changes between benign and malignant mammary tumours, we have analysed matched normal and adenoma-associated stroma (AAS) from 13 canine mammary adenomas and compared them to previous data from 15 canine mCA. Our analyses reveal distinct stromal reprogramming even in small benign tumours. While similarities between AAS and CAS exist, the stromal signature clearly distinguished adenomas from mCA. The distinction between AAS and CAS is further substantiated by differential enrichment in several hallmark signalling pathways as well as differential abundance in cellular composition. Finally, we identify COL11A1, VIT, CD74, HLA-DRA, STRA6, IGFBP4, PIGR, and TNIP1 as strongly discriminatory stromal genes between adenoma and mCA, and demonstrate their prognostic value for human breast cancer. Given the relevance of canine CAS as a model for the human disease, our approach identifies disease-modulating stromal components with implications for both human and canine breast cancer.

Biphasic Feline Mammary Carcinomas Including Carcinoma and Malignant Myoepithelioma

Feline mammary tumors are usually malignant and aggressive carcinomas. Most cases are simple monophasic carcinomas (1 epithelial population), and additional phenotyping is usually not needed. In this study, we describe 10 malignant mammary tumors from 9 female cats that had unusual histomorphology: they appeared biphasic, with 2 distinct cell populations. Initially, they were morphologically diagnosed as either carcinosarcoma (1/10) or malignant pleomorphic tumor (9/10) of the mammary gland, as the latter did not match any previously described histological subtype. Immunohistochemistry (IHC) was performed for pancytokeratin, cytokeratins 8 and 18, cytokeratin 14, cytokeratins 5 and 6, vimentin, p63, calponin, alpha-smooth muscle actin, Ki-67, ERBB2, estrogen receptor alpha, and progesterone receptor. In 7 of 10 cases, the biphasic nature was confirmed and, on the basis of the IHC results, they were classified as carcinoma and malignant myoepithelioma (4/10), ductal carcinoma (1/10), and carcinosarcoma (2/10). The other 3 of 10 cases were monophasic based on IHC. In the cases of carcinoma and malignant myoepithelioma, the malignant myoepithelial cells were 100% positive for vimentin (4/4) and variably positive for p63, calponin, and cytokeratins (4/4). These findings show that, although rare, biphasic mammary carcinomas do occur in cats. In dogs and humans, tumors composed of malignant epithelial and myoepithelial cells have a less aggressive behavior than certain simple carcinomas, and therefore, their identification might also be clinically significant in the cat.

Clinical and Ultrasonographic Aspects of Benign and Malignant Mammary Tumors in Female Dogs

Background: Mammary neoplasms in dogs are commonly observed in veterinary clinical routine, most of which being malignant. Hormonal stimulation, endogenous or exogenous, may possibly influence its development. In addition to clinical evaluation, ultrasound analysis can provide information about the characteristics of breast lumps. The association between clinical-epidemiological and pathological data is important for diagnosis. Therefore, given the importance of this pathology for the health of affected dogs, we aimed to evaluate the clinical and ultrasound alterations, along with the factors associated with the development of benign and malignant mammary neoplasms in female dogs. Materials, Methods &amp; Results: We examined 47 samples from the mammary tumors of 35 female dogs at the Small Animal Clinic of the Veterinary Hospital (HV) of the Santa Cruz State University (UESC). The dogs underwent a complete clinical examination, with clinical staging, via TNM classification, followed by hematological, biochemical, radiological and ultrasound, abdominal, and breast exams. Breast ultrasound examination was used to evaluate the shape parameters such as, limits, margins or contour, ecotexture, echogenicity, hyperechoic halo, posterior acoustic shading, surrounding changes, and nodule components. These criteria were associated with the histopathological classification of neoplasms. Epidemiological data was studied through an adapted questionnaire containing information on risk factors associated with breast cancer. The same questionnaire was applied to tutors of 19, age-matched, female dogs with no history of breast cancer. The results revealed that most female dogs with neoplasia were over eight years of age, with no specific breed and were not castrated, and 31.4% of them had already been administered with contraceptives during the reproductive period. Ovariohysterectomy acted as a protective factor (OR 0.06) to the development of breast tumors, while contraceptive use was considered as a risk factor (OR 6.99). The average time reported between tumor perception and clinical care was 13.2 months. The caudal and inguinal abdominal breasts were the most affected. Among the samples evaluated, 76.6% were malignant, with mixed tumor carcinoma being the most frequent type and 69.4% graded in grade I. Nodules classified as malignant showed the largest diameter (P &lt; 0.05). Breast ultrasound results revealed that tumors with heterogeneous echotextures and mixed components were associated with malignancy (P &lt; 0.05). Discussion: The fact that the average age of female dogs with breast cancer was over eight years of age corroborates the literature. Considering that a greater age would mean a longer exposure to the carcinogenic initiators responsible for mutations, and to promoters, such as hormonal changes. Contraceptives increase the risk of breast lumps, while reduce that of ovariohysterectomy, in female dogs, even when performed after the second heat. The size of the nodules and ultrasound criteria related to echotexture and the type of component of the neoplasia may be used as prognostic parameters of female breast nodules. Additionally, most nodules evaluated in this study were malignant (mixed tumor carcinoma was the most common subtype), possibly due to the owners' delay in seeking veterinary care after tumor observation. Although malignant, most nodules presented with a low histopathological grading.Background: Mammary neoplasms in dogs are commonly observed in veterinary clinicalroutine, most of which being malignant. Hormonal stimulation, endogenous or exogenous,may possibly influence its development. In addition to clinical evaluation, ultrasound analysiscan provide information about the characteristics of breast lumps. The association betweenclinical-epidemiological and pathological data is important for diagnosis. Therefore, given theimportance of this pathology for the health of affected dogs, we aimed to evaluate the clinicaland ultrasound alterations, along with the factors associated with the development of benignand malignant mammary neoplasms in female dogs.Materials, Methods &amp;amp; Results: We examined 47

Identification of p53 gene alterations in canine mammary tumours using polymerase chain reaction and direct sequence analysis

Mammary tumours are mentioned as the most common tumours in female dogs and approximately half of them are detected malignant. p53 gene mutations are demonstrated to be the most common genetic alteration in canine mammary tumours. The present study was conducted to evaluate exon-1 of p53 gene mutations in tissue samples of canine mammary tumours by PCR and direct sequence analysis. After histopathological confirmation of the tissue sections by haematoxylin and eosin staining (10/26), deparaffinised samples were used for DNA extraction by silica gel method. Subsequently, p53 exon 1 was amplified through PCR assay using specific oligo nucleotide primers designed according to the canine DNA sequence available online. Microscopically, 10 out of 26 suspected tissue samples were recognised as malignant mammary gland tumours with various grades of malignancy. Surprisingly, one insertion of mutation was found in exon 1 of all examined samples corresponding to a sequence comprising 27 amino acids, between amino acids 30 to 57 in the p53 protein. Taken together, it seems that alteration of exon 1 p53 gene may lead to malignancy beha­viour, poor prognosis and short survival time in dogs with mammary carcinomas.

Abdominal ultrasonographic findings in dogs with mammary tumors: association with tumor characteristics and survival

Abdominal ultrasound is frequently performed as part of initial staging of canine mammary tumors. The aims of this study were to describe abdominal ultrasonographic findings present at diagnosis of mammary tumors in dogs and to assess their value for associations with histopathological results and treatment outcome. The medical records of 201 female dogs diagnosed with mammary tumors that had abdominal ultrasonography performed as part of their initial clinical evaluation were reviewed. Histopathological diagnosis of malignant mammary tumors was obtained in most cases (86%; 172 cases). One hundred and fifty-seven animals (78%) had some ultrasonographic abnormality. A total of 338 ultrasonographic changes were identified, and the spleen was the organ with the highest frequency of ultrasonographic changes (20%), followed by the uterus (18%) and the liver (16%). Abdominal ultrasonography abnormalities were not associated with mammary tumor malignancy (OR = 1.7; P = 0.27). Ultrasonographic evidence of enlarged intraabdominal lymph nodes was associated with metastases in surface regional lymph nodes (P < 0.05). Dogs with regional metastasis were more likely to present some abdominal ultrasonographic change (OR = 6.1; P = 0.04). Animals with ultrasonographic evidence of intraabdominal lymph node enlargement presented shorter survival after mastectomy. Abdominal ultrasonographic abnormalities are common in dogs with mammary tumors and are not associated with malignancy. However, the presence of intraabdominal lymph node enlargement identified by ultrasound may be associated with regional metastasis and shorter survival rates.