Malignant Canine Mammary Tumors Research Articles

Detection of PIK3CA hotspot mutations in canine mammary tumors using droplet digital PCR: tissue validation and liquid biopsy feasibility

Domestic dogs (Canis lupus familiaris) serve as valuable translational models for human cancer research due to their biological similarities. Canine mammary tumors (CMTs), frequently diagnosed in female dogs, share various characteristics with human breast cancers. This study investigates the PIK3CA (H1047R) mutation in CMTs using droplet digital PCR (ddPCR) and explores the potential of liquid biopsy for non-invasive detection. We analyzed 80 formalin-fixed, paraffin-embedded (FFPE) CMT tissue samples and compared ddPCR results with next-generation sequencing (NGS) data, achieving high concordance. Plasma and serum samples were also assessed for mutation concordance with tissue results. Our findings indicate a higher frequency of the PIK3CA (H1047R) mutations in benign and grade I malignant CMTs compared to more aggressive malignancies. The ddPCR assay demonstrated high sensitivity and specificity, with plasma testing showing 78.6% sensitivity and 87.5% specificity, and serum testing showing 66.7% sensitivity and 90.0% specificity. These results highlight the viability of liquid biopsy as a minimally invasive method for monitoring PIK3CA mutations in canine patients. The study suggests that liquid biopsy techniques hold significant promise for improving the early detection and monitoring of canine cancers, warranting further research to refine these methods and explore their applications in canine cancer diagnostics and treatment.

TGFβ in malignant canine mammary tumors: relation with angiogenesis, immunologic markers and prognostic role

Transforming growth factor-β (TGFβ) and FoxP3 regulatory T cells (Treg) are involved in human breast carcinogenesis. This topic is not well documented in canine mammary tumors (CMT). In this work, the tumoral TGFβ expression was assessed by immunohistochemistry in 67 malignant CMT and its correlation to previously determined FoxP3, VEGF, and CD31 markers and other clinicopathologic parameters was evaluated. The high levels of TGFβ were statistically significantly associated with skin ulceration, tumor necrosis, high histological grade of malignancy (HGM), presence of neoplastic intravascular emboli and presence of lymph node metastases. The observed levels of TGFβ were positively correlated with intratumoral FoxP3 (strong correlation), VEGF (weak correlation) and CD31 (moderate correlation). Tumors that presented a concurrent high expression of TGFβ/FoxP3, TGFβ/VEGF, and TGFβ/CD31 markers were statistically significantly associated with parameters of tumor malignancy (high HGM, presence of vascular emboli and nodal metastasis). Additionally, shorter overall survival (OS) time was statistically significantly associated with tumors with an abundant TGFβ expression and with concurrent high expression of TGFβ/FoxP3, TGFβ/VEGF, and TGFβ/CD31. The presence of lymph node metastasis increased 11 times the risk of disease-related death, arising as an independent predictor of poor prognosis in the multivariable analysis. In conclusion, TGFβ and Treg cells seem involved in tumor progression emerging as potential therapeutic targets for future immunotherapy studies.

Evaluation and Comparison of Different Surgico-chemotherapeutic Regimens for the Treatment of Malignant Canine Mammary Tumours

Background: Cancer has been regarded as the most dreaded disease for both human and animals because of its incurability. Canine mammary tumours are common and major problem encountered by academicians and practicing veterinarians. As a result, the current study was conducted to examine the effectiveness and overall impact of different surgico-chemotherapeutic approaches in treating malignant mammary tumors in dogs. Methods: The investigation involved 18 clinical instances of mammary tumors in different breeds, regardless of age or sex and were divided into three groups of six animals each. In Group S, only surgical excision of tumour was performed while animals of Group SD and Group SV were treated with surgical excision followed by administration of Doxorubicin (30 mg/m2) BSA and Vincristine sulphate (0.025 mg/kg) intravenously alongwith DNS at 7th and 14th post-operative days respectively. Different physiological and haemato-bichemical parameters i.e. Hb, PCV, TLC, TPC, DLC, Serum glucose, TSP, SUN, SC, ALT, AST and ALP were recorded preoperatively, postoperatively and following chemotherapy at 10th, 30th and 60th day intervals. Result: The present study showed transient change in the physiological, haematological and biochemical profiles following surgery and chemotherapy treatment. The histopathological analysis showed more cases of adenocarinoma which were followed by mixed carcinoma. Surgery combined with chemotherapy (doxorubicin and vincristine) resulted in minimal to no reoccurrence with few adverse reactions including inappetance, vomition, anaemia and alopecia. It was concluded that the most effective treatment for malignant mammary tumors in dogs is surgical excision followed by sequential vincristine therapy, which efficiently regresses the tumor without causing relapse.

Investigation of efficacy of two different chemotherapy protocols used in neoadjuvant chemotherapy in clinical stages II-IV canine malignant mammary tumours.

The first aim of this study is to demonstrate the clinical efficacy and reliability of two different neoadjuvant chemotherapy (NAC) protocols consisting of doxorubicin/cyclophosphamide (AC) and paclitaxel in dogs with clinical stages II-IV canine malignant mammary tumours (CMTs). Secondly, to determine the Luminal A, Luminal B, HER2-positive and triple-negative molecular subtypes and their value in predicting clinical response to NAC in biopsy samples, and thirdly, to reveal the changes in Ki-67, human epidermal growth factor receptor type 2 (HER2), oestrogen receptor (ER), and progesterone receptor (PgR) expression levels induced by NAC. Thirty dogs with clinical stages II-IV CMTs (T1-3N0-1M0) according to the modified TNM system were included in the study. Dogs in group-1 (n = 15) AC combination and dogs in group-2 (n = 15) were administered paclitaxel. Partial response (PR) was the most common clinical response in both treatment groups (66.66% and 86.66%, respectively). There was no difference between the groups regarding clinical response parameters (p = .001). The rate of treatment responders was higher than the rate of non-responders in both groups (p < .001). The adverse effects observed in both groups were mostly limited to grades 1 and 2 and all were easy to manage. The most frequently detected molecular subtype was Luminal A (59.25%). Complete response (CR) was achieved in 33.33% of dogs with triple-negative CMT in the AC group and 14.29% of the Luminal A subtype in the paclitaxel group. Alterations in Ki-67, HER2, ER, and PgR expressions after chemotherapy were not statistically significant (p > .05). As a result, we have shown that these neoadjuvant chemotherapy protocols are effective and safe alternative treatment options for CMTs.

Risk factor analysis and clinicopathological characteristics of female dogs with mammary tumours from a single-center retrospective study in Poland

This is a comprehensive retrospective study to characterize female dogs with canine mammary tumors (CMTs) using a dataset retrieved from the archives of the Division of Animal Pathology, Institute of Veterinary Medicine in Warsaw, and to identify prognostic factors. Clinical and histopathological data of 1447 dogs with CMTs were included. Malignant tumours were found in 83.3% (n = 1206), benign tumours in 11.7% (n = 169), and non-neoplastic lesions in 5.0% (n = 72) of dogs. Dogs most often had grade II carcinomas (38.2%, 215/562) of a single histological subtype (88.5%, 1281/1447), mostly simple carcinoma (35.3%, 510/1447). Dogs with a median age of 10 years significantly often had larger (≥ 3 cm) and malignant CMTs, whereas intact females had smaller tumours (median size 2.0 cm). However, the threshold value for the age of the dog in the differentiation of malignant and non-neoplastic/benign masses could not be determined. Most females were hormonally active (76.4%, 372/487). Hormonally active dogs significantly more often had multiple tumours. Multiple tumours were significantly smaller (median 2.5 cm) than single ones. Among pedigree dogs, small-breed dogs were mostly recorded (43%, 428/1006). Twelve breeds had an increased risk of CMTs, regardless of tumour behaviour, compared with the theoretical distribution of pedigree dogs in Poland. Four breeds were often affected only by malignant and other four breeds only by non-neoplastic/benign CMT. Large-breed dogs were significantly younger and affected by larger CMT (median 4 cm) compared with small- and medium-breed dogs. Ninety dogs with a malignant CMT and complete records were included in the full analysis of CMT-specific survival (CMT-SS) with a median follow-up time of 20.0 months. We showed that the timing of ovariohysterectomy in relation to mastectomy was significantly associated with grade, CMT-SS, and CMT-related death. We indicated the low diagnostic accuracy of palpation of regional lymph nodes (RLN) in the prediction of their metastatic involvement. By multivariable analysis, dogs with neoplastic emboli, tumour ulceration, and simple or complex carcinoma had a significantly higher risk of local recurrence. Tumour size > 3 cm was as a strong independent predictor of lung metastases. Compared with dogs with an easily separated localized tumour, dogs with a multiple/diffuse malignant CMT pattern had a fivefold higher risk of death. The risk of death was significantly higher in the presence of neoplastic emboli (~ fivefold) and tumour ulceration (~ fourfold). Furthermore, the presence of neoplastic emboli and large tumour size were independent predictors of CMT-related death.

Machine learning determines stemness associated with simple and basal-like canine mammary carcinomas

Simple and complex carcinomas are the most common type of malignant Canine Mammary Tumors (CMTs), with simple carcinomas exhibiting aggressive behavior and poorer prognostic. Stemness is an ability associated with cancer initiation, malignancy, and therapeutic resistance, but is still few elucidated in canine mammary tumor subtypes. Here, we first validated, using CMT samples, a previously published canine one-class logistic regression machine learning algorithm (OCLR) to predict stemness (mRNAsi) in canine cancer cells. Then, using the canine mRNAsi, we observed that simple carcinomas exhibit higher stemness than complex carcinomas and other histological subtypes. Also, we confirmed that stemness is higher and associated with basal-like CMTs and with NMF2 metagene signature, a tumor-specific DNA-repair metagene signature. Using correlation analysis, we selected the top 50 genes correlated with higher stemness, and the top 50 genes correlated with lower stemness and further performed a gene set enrichment analysis to observe the biological processes enriched for these genes. Finally, we suggested two promise stemness-associated targets in CMTs, POLA2 and APEX1, especially in simple carcinomas. Thus, our work elucidates stemness as a potential mechanism behind the aggressiveness and development of canine mammary tumors, especially in simple carcinomas, describing evidence of a promising strategy to target this disease.

Immunohistochemical Study of MMP-9 Expression in Canine Malignant Mammary Tumors

A class of proteinases called matrix metalloproteinases (MMPs) is involved in the breakdown of the extracellular matrix (ECM) as well as other biological activities. Due to their capacity to degrade the matrix, they were considered in the invasion and metastasis of tumors. The primary goal of this study is to evaluate the association between MMP-9 expression in tumor-adjacent stromal cells and malignant mammary tumors in dogs, as well as the potential use of this protease as a predictive tumor marker. During surgery and necropsy, 32 canine malignant mammary gland tumors were discovered. For histopathological grading of tumors, the Pena et al. [1], approach and the Goldsmith et al. [2], method were employed. Utilizing immunohistochemical labeling, epithelial and stromal cell MMP-9 expression was identified. The level of tumor malignancy was associated with the degree of MMP-9 expression in stromal cells. In addition, stromal cells had much greater levels of MMP-9 expression. The expression of MMP-9 was well distributed and intense in all grade II and III carcinomas as well as poorly differentiated carcinosarcomas. High histologic grade tumors have greater levels of MMP-9 expression, according to semiquantitative examination of the gene’s expression. This finding lends credence to the concept that MMP-9 functions as an ECM component in tumor invasion and metastasis.

Advancing canine mammary tumor diagnostics: Unraveling the diagnostic potential of Cytokeratin 19 through droplet digital PCR analysis

Cytokeratin 19 (CK19) is a complex intracytoplasmic cytoskeletal protein primarily localized in the ducts of the mammary gland and skin epithelial cells. In humans, the expression of CK19 gene within circulating tumor cells (CTCs) extracted from blood samples of breast cancer patients reflects tumor cell activity, offering valuable insights for predicting early metastatic relapse or monitoring treatment effectiveness. However, knowledge of serum tumor markers is limited in veterinary oncology. Recently, droplet digital PCR (ddPCR), has been employed to explore rare target genes due to its heightened sensitivity and accuracy as a novel molecular diagnostic tool. The objectives of this study were to investigate the expression of the CK19 mRNA in CTCs, non-neoplastic mammary tissues, and both benign and malignant canine mammary tumors (CMTs) through ddPCR analysis. In Study I, we optimized the discard volume for blood samples to reduce CK19 contamination from skin epithelial cells post-venipuncture. The results revealed that discarding the initial 3 mL of blood was adequate and effective in eliminating CK19 mRNA contamination. In Study II, after the removal of the initial 3 mL of blood, we investigated CK19 mRNA-positive CTCs in the peripheral blood of normal healthy dogs, including those with benign and malignant CMTs. Intriguingly, CK19 mRNA was undetectable in all blood samples. The expression of CK19 mRNA in mammary tissues was investigated in Study III. The copy number (CN) ratios of the CK19 gene in non-neoplastic mammary tissues (14.77 ± 14.65) were significantly higher (P < 0.05) than those in benign (4.23 ± 3.35) and malignant groups (6.56 ± 5.64). Notably, no difference was observed between the benign and malignant groups. In conclusion, CK19 mRNA appeared unlikely to be a suitable candidate as a biomarker in the peripheral blood of CMTs, while the CN ratio in mammary tissues could serve as a potential discriminator between non-neoplastic and CMT groups, complementing the gold standard of histopathological examination.

Vascular Endothelial Growth Factor A (VEGF-A) concentrations in canine malignant mammary tumours with and without metastases to regional lymph nodes

Vascular Endothelial Growth Factor A (VEGF-A) concentrations in canine malignant mammary tumours with and without metastases to regional lymph nodes

YRNA and tRNA fragments can differentiate benign from malignant canine mammary gland tumors

Mammary gland tumors (MGT) are the most common tumors in sexually intact female dogs. The functional regulation of miRNAs, a type of noncoding RNAs (ncRNAs), in canine MGT has been extensively investigated. However, the expression of other ncRNAs, such as YRNAs and transfer RNA–derived fragments (tRFs) in canine MGT is unknown. We investigated ncRNAs other than miRNAs from our small RNA project (PRJNA716131) in different canine MGT histologic subtypes. This study included benign tumors (benign mixed tumor, complex adenoma) and malignant tumors (carcinoma in benign tumor and carcinoma with metastasis) samples. Aberrantly expressed ncRNAs were examined by comparisons among MGT subtypes. The relative expression trends were validated in canine MGT tissues, plasma, extracellular vesicles, and MGT cell lines using quantitative reverse transcription PCR. Three aberrantly expressed ncRNAs were identified by comparisons among MGT subtypes. YRNA and tRNA-Gly-GCC distinguished benign mixed tumor from other MGT histologic subtypes, while tRNA-Val differentiated complex adenoma, carcinoma in benign tumors, and carcinoma with metastasis. The ROC curve of the three ncRNAs showed they might be potential biomarkers to discriminate malignant from benign MGT. YRNA and tRFs expression levels were decreased in metastatic compared with primary canine MGT cell lines. To the best of our knowledge, this is the first investigation of YRNA and tRFs in canine MGT. The three identified ncRNAs may be biomarkers for differentiating MGT histologic subtypes. Suggested Reviewers: Powered by Editorial Manager® and ProduXion Manager® from Aries Systems Corporatio

Immunoexpression of the cannabinoid CB2 receptor protein in canine benign and malignant mammary tumours

Immunoexpression of the cannabinoid CB2 receptor protein in canine benign and malignant mammary tumours

Epigenetics in Canine Mammary Tumors: Upregulation of miR-18a and miR-18b Oncogenes Is Associated with Decreased ERS1 Target mRNA Expression and ERα Immunoexpression in Highly Proliferating Carcinomas

The expression of miRNAs is one of the main epigenetic mechanisms responsible for the regulation of gene expression in mammals, and in cancer, miRNAs participate by regulating the expression of protein-coding cancer-associated genes. In canine mammary tumors (CMTs), the ESR1 gene encodes for ERα, and represents a major target gene for miR-18a and miR-18b, previously found to be overexpressed in mammary carcinomas. A loss in ERα expression in CMTs is commonly associated with poor prognosis, and it is noteworthy that the downregulation of the ESR1 would appear to be more epigenetic than genetic in nature. In this study, the expression of ESR1 mRNA in formalin-fixed, paraffin-embedded (FFPE) canine mammary tumors (CMTs) was evaluated and compared with the expression levels of miR18a and miR18b, both assessed via RT-qPCR. Furthermore, the possible correlation between the miRNA expression data and the immunohistochemical prognostic factors (ERα immunoexpression; Ki67 proliferative index) was explored. A total of twenty-six FFPE mammary samples were used, including 22 CMTs (7 benign; 15 malignant) and four control samples (three normal mammary glands and one case of lobular hyperplasia). The obtained results demonstrate that miR-18a and miR-18b are upregulated in malignant CMTs, negatively correlating with the expression of target ESR1 mRNA. Of note, the upregulation of miRNAs strictly reflects the progressive loss of ERα immunoexpression and increased tumor cell proliferation as measured using the Ki67 index. The results suggest a central role of miR-18a and miR-18b in the pathophysiology of canine mammary tumors as potential epigenetic mechanisms involved in ERα downregulation. Moreover, as miRNA expression reflects ERα protein status and a high proliferative index, miR-18a and miR-18b may represent promising biomarkers with prognostic value. More detailed investigations on a larger number of cases are needed to better understand the influence of these miRNAs in canine mammary tumors.

Expression Pattern of SLIT 2 Genes Discriminates Malignant from Benign Canine Mammary Tumours

The study was conducted to investigate the expression of SLIT2 gene in representative tissue samples of mammary tumours of female dogs (n=10) with the age between 5.0 and 16.5 years brought to Veterinary Clinical Complex, LUVAS, Hisar. Excisional biopsy was taken after surgery of these cases. Histopathological studies confirmed four cases of adenocarcinoma, three cases of benign mixed tumours and two of myoepithelioma and one case of adenoma. Amplification plots and dissociation curves of RT-qPCR were indicative of specific amplification of the canine SLIT 2 gene in all the samples. Further, amplification product of a 160 bp SLIT 2 gene and 120 bp β-actin gene in all the RT-qPCR tubes were visible by 2% agarose gel electrophoresis. Out of 10 tissue samples examined, transcript levels of SLIT 2 genes in all different samples ranged from 1.90 to 84.98. Conclusively, the expression of SLIT 2 genes increased in all canine malignant mammary gland tumours, thereby showing its association with tumour malignancy.

Evaluation of Some Systemic Inflammatory Biomarkers in Canine Malignant Mammary Tumors

Evaluation of Some Systemic Inflammatory Biomarkers in Canine Malignant Mammary Tumors

Evaluation of tumour associated macrophages in different histopathological types and grades of canine mammary tumours

Canine mammary tumours (CMTs) are the second highest reported tumours in female dogs, following skin tumours. Human breast cancers (HBCs) and CMTs share common clinical and molecular features and hence, CMTs are considered as ideal models to study the different aspects of HBC. The study utilised samples from 25 CMT suspected cases presented to University Veterinary Hospitals in Thrissur district from December 2020 to October 2021. The tumour samples were analysed histopathologically and the lesions were classified. Among the 25 cases, one was identified as ductal hyperplasia, one as a benign myxoma and all the others were found to be malignant neoplasms. Malignant tumours were further categorised into different histotypes.Histological Malignancy Grading (HMG) was also done in 23 malignant CMTs and 21.74 percent were found to be of grade I, 47.83 per cent were grade II and 30.43 per cent were grade III. Majority of the malignant tumours were simple carcinomas which comprised tubulopapillary, ductal, cribriform, solid and comedocarcinomas. Highly aggressive tumours like cribriform, solid, comedo and inflammatory carcinomas belonged to higher grades, either II or III. Infiltration of tumour associated macrophages (TAMs) was studied in different histotypes and grades of CMTs. It was identified that malignant high grade CMTs had greater TAM infiltration and hence, with further validation TAMs could be effectively used in predicting prognosis and also as a therapeutic target.

Na+/K+-ATPase and bone morphogenetic protein-2 expressions in parenchymal and microenvironmental cells of canine mammary tumours.

The most common canine tumour is mammary tumour, which resembles breast cancer in humans. Microenvironment is a crucial factor in the formation of breast cancers. In order to distinguish between benign and malignant canine mammary tumours, this study looked at the immunohistochemical expression of Na+/K+-ATPase and bone morphogenetic protein-2 (BMP-2) in tumour and microenvironmental cells. The aim of this study was to evaluate the expression of Na+/K+-ATPase and BMP-2 in canine mammary tumours and their relationship with malignancy. In this investigation, 10 normal breast tissues were used as controls, and 28 benign and 46 malignant mammary tumours were taken from the archives of the Department of Pathology. The findings showed that malignant tumours expressed more Na+/K+-ATPase and BMP-2 than did normal breast tissue. Both markers had a negative or slight expression in benign tumours, whereas they considerably increased in malignant tumours. Both tumour parenchymal and microenvironmental cells in malignancies expressed Na+/K+-ATPase and BMP-2. Na+/K+-ATPase expression was observed to be more prominent in cells when compared to BMP-2. These findings also suggest that Na+/K+-ATPase and BMP-2 could be employed in the future to help diagnose canine and possibly human breast cancers earlier or as possible targets for treatment.

Molecular phenotyping of malignant canine mammary tumours: Detection of high-risk group and its relationship with clinicomolecular characteristics.

Canine mammary gland tumours (CMTs) constitute the most common cancer in female dogs and comprise approximately 50% of all canine cancers. With the advent of high-throughput technologies such as microarray and next-generation sequencing, the molecular phenotyping (classification) of various cancers has been extensively developed. The present study used a canine RNA-sequencing dataset, namely GSE119810, to classify 113 malignant CMTs and 64 matched normal samples via an unsupervised hierarchical algorithm with a view to evaluating the association between the resulting subtypes (clusters) (n= 4) and clinical and molecular characteristics. Finally, a molecular classifier was developed, and it detected 1 high-risk molecular subtype in the training dataset (GSE119810) and 2 independent validation datasets (GSE20718 and GSE22516). Our results revealed four molecular subtypes (C2-C5) in malignant CMTs. Furthermore, the normal samples constituted a distinct group in the clustering analysis. Marked significant associations were observed between the molecular subtypes (especially C5) and clinical/molecular features, including positive lymphatic invasion, high tumour grades, histopathology diagnoses, short survival and high TP53 mutation rates (ps<.05). The high-risk subtype (C5) was further characterized through the development of a cell cycle-based gene signature, which comprised 37 proliferation-related genes according to the support vector machine algorithm. This signature identified the high-risk group in both training and validation datasets (ps<.001). In the validation analysis, our potential classifier robustly predicted patients with positive lymphatic invasion, metastases and short survival.

Plasma interleukin-1α and interleukin-8 in female dogs with non-metastatic and metastatic malignant mammary gland tumours.

In this study plasma concentrations of IL-1α and IL-8 in 29 female dogs with malignant mammary gland tumours (19 without metastasis and 10 with metastasis) and in 10 healthy control animals were determined. Concentrations of IL-1α and IL-8 were analysed using a specific canine ELISA assay. Mean plasma concentrations of IL-1α and IL-8 were significantly higher (p⟨0.05) in female dogs with both non-metastatic and metastatic malignant tumours compared to the healthy animals. The concentrations of both tested cytokines were significantly increased (p⟨0.05) in the dogs with metastasis. In female dogs with mammary carcinomas, the plasma concentration of IL-1α was significantly higher (p⟨0.05) in the animals with grade 3 tumours compared to the dogs with grade 1 tumours. The concentration of IL-8 was significantly higher (p⟨0.05) in the dogs with grade 3 tumours compared to that found in the animals with grade 1 and grade 2 tumours. A moderate correlation (r=0.433) was found between IL-1α and IL-8 concentrations in the female dogs. These findings suggest that increased malignancy and invasiveness of canine mammary tumours is associated with an increased production of IL-1α and IL-8 in the tumour microenvironment, which, in turn, leads to an increase in their circulating levels. This may indicate that circulating levels of the cytokines investigated could be considered as diagnostic and prognostic markers in canine malignant mammary tumours. However, further studies in this fields are needed.

Significância prognóstica dos imunofenótipos em tumores mamários malignos caninos

ABSTRACT Canine malignant mammary neoplasms (CMMN) exhibit behavioral variability with the patient survival time depending on several prognostic factors. In the present study, 134 CMMN were selected and different immunophenotypes and their associations with clinical and pathological parameters were identified. The tumors were classified as follows: 46% of luminal B HER2-, 34% of luminal A, 13% of triple-negative, and 7% of luminal B HER2+. Shorter specific survival time were associated with larger tumor sizes (&gt;3.0 cm, HR=1.94; P=0.0209), lymph node metastasis or distant metastasis (HR= 2.82; P &lt;.0001), more aggressive histological types (HR= 7.15, P&lt;0.0001), higher histological grades (HR= 12.97 P=0.011), angiolymphatic invasion (HR=4.68, P&lt;0.0001) and luminal B HER2 - (HR= 3.27, P&lt;0.0001) and luminal B HER2 + (HR= 7.14 P&lt;0.0001) immunophenotypes. In patients with lymph nodal metastasis, shorter survival times were associated with luminal immunophenotype B HER2 + (P=0.003). However, in patients without metastasis, an increased risk of death was associated with the aggressive histological type. In conclusion, the classification in our study allowed us to identify subtypes with different prognoses in canine malignant mammary tumors. Factors such as clinical stage, histological type, luminal B HER2+ subtype, and angiolymphatic invasion were the most important prognostic factors.

Histomorphological stratification of stromal types associated with canine mammary tumours

Cancer associated stroma (CAS) consists mainly of a cellular fraction comprising fibroblasts, myofibroblasts, inflammatory cells, immune cells, endothelial cells, adipocytes and extracellular matrix (ECM). Recent reports have shown that the cancer stroma including the cellular fraction and ECM undergo considerable reprogramming during the process of tumourigenesis. Though there are some studies on CAS of human breast cancers (HBCs), similar studies are very much limited in canine mammary tumours. Hence, the present study was undertaken to classify the stromal types associated with malignant canine mammary tumours(CMTs). The excisional biopsy samples from 50 numbers of CMTs presented to Kerala Veterinary and Animal Sciences University hospitals at Mannuthy and Kokkalai during the period from November 2019 to December 2021 formed the study material. Histopathological stratification of cancer stroma was done using qualitative evaluation based on the stromal characteristics suggested for HBCs. Accordingly the cancer associated stroma in different tumour samples were classified as fibrotic stroma, inflammatory stroma and mixed stroma. Fibrotic stroma was further classified as mature/ sclerotic, intermediate and immature/desmoplastic types. The grade of tumours in relation to the type of stroma was also analysed and it was identified that with the exception of sclerotic stroma, all other stromal types could be observed in higher grades of CMTs. Desmoplastic and inflammatory stroma were predominantly seen associated with Grade II and Grade III tumours.