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Programmed death ligand 1 (PD-L1) is an immune checkpoint molecule that plays a crucial role in regulating antitumor immune responses. Canine mammary carcinomas (CMCs) are common tumors of dogs. Despite extensive studies on the heterogeneity of CMCs, there is still a lack of effective precision therapies for the treatment of CMCs. In this study, we aimed to investigate the correlation between PD-L1 mRNA and protein expression in CMCs and explore its association with histopathological grade and molecular markers, including the estrogen receptor, epidermal growth factor receptor 2, and cytokeratin 5/6 (CK5/6). Formalin-fixed paraffin-embedded samples were evaluated for PD-L1 mRNA expression using RNA in situ hybridization and PD-L1 protein expression using immunohistochemistry. We observed no substantial correlation between PD-L1 mRNA and protein expression in CMCs; however, PD-L1 mRNA levels were significantly higher in grade 3 than in grade 1 tumors (P = .001). In addition, we observed a positive correlation between PD-L1 protein expression and CK5/6 expression in CMCs (P = .032). These findings suggest that PD-L1 expression in CMCs is heterogeneous and may be regulated post-transcriptionally. Further studies are needed to explore the prognostic and therapeutic implications of PD-L1 expression in different molecular subtypes of CMCs and their potential as predictive biomarkers for immunotherapy.
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Antígeno B7-H1 , Biomarcadores de Tumor , Enfermedades de los Perros , Neoplasias Mamarias Animales , ARN Mensajero , Animales , Perros , Antígeno B7-H1/metabolismo , Antígeno B7-H1/genética , Femenino , Neoplasias Mamarias Animales/metabolismo , Neoplasias Mamarias Animales/patología , Neoplasias Mamarias Animales/genética , Enfermedades de los Perros/patología , Enfermedades de los Perros/metabolismo , Enfermedades de los Perros/genética , ARN Mensajero/metabolismo , ARN Mensajero/genética , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Inmunohistoquímica/veterinaria , Regulación Neoplásica de la Expresión GénicaRESUMEN
PURPOSE: Mammary gland tumors are the most common neoplastic diseases in elderly female dogs, about 50% of which are considered to be malignant. Canine mammary tumors are similar to human breast cancers in many respects, so canine mammary tumors are frequently studied alongside human breast cancer. This article mentioned KI-67, HER-2, COX-2, BRCA1, BRCA2, P53, CA15-3, MicroRNA, Top2α and so on. All these markers are expected to have an important role in the clinic. METHODS: Existing markers of canine mammary carcinoma are reviewed, and the expression of each marker and its diagnostic role for this tumor are described in detail. RESULTS: This article introduced several effective markers of canine mammary tumors, among them, antigen KI-67 (KI-67), human epidermal growth factor receptor 2 (HER-2), cyclooxygenase 2 (COX-2) are promising and can be detected in both serum and tissue samples. Breast cancer caused by mutations in the breast cancer 1 gene (BRCA1) and breast cancer 2 gene (BRCA2) is also a hot topic of research. In addition to the above symbols, tumor protein p53 (p53), cancer antigen15-3 (CA15-3), MicroRNA (miRNA), topoisomerase πα (Top2α), proliferating cell nuclear antigen (PCNA), epidermal growth factor receptor (EGFR) and E-cadherin will also be involved in this paper. We will also mention Mammaglobin, which has been rarely reported so far.
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Neoplasias de la Mama , Carcinoma , Enfermedades de los Perros , Neoplasias Mamarias Animales , MicroARNs , Humanos , Animales , Perros , Femenino , Anciano , Antígeno Ki-67/metabolismo , Proteína p53 Supresora de Tumor/genética , Ciclooxigenasa 2/genética , Ciclooxigenasa 2/metabolismo , Neoplasias Mamarias Animales/genética , Neoplasias Mamarias Animales/metabolismo , Carcinoma/genética , Neoplasias de la Mama/genética , MicroARNs/genética , Enfermedades de los Perros/diagnóstico , Enfermedades de los Perros/genética , Enfermedades de los Perros/metabolismo , Regulación Neoplásica de la Expresión GénicaRESUMEN
As one of the most common malignancies in female dogs, no drugs have been developed specifically for the treatment of canine mammary carcinoma. In our previous study, a series of diterpenoid alkaloids derivatives were synthesized and exhibited good anti-proliferative activity in vitro against both normal and adriamycin-resistant human breast cancer cells lines. In this study, a series of structurally diverse aconitine-type alkaloids derivatives were also synthesized basing on the minimal modification principle, by modifying on A-ring, C-ring, D-ring, N-atom or salt formation on aconitine skeleton. Their anti-proliferative effects and mechanism on canine mammary cancer cells were investigated, exhibiting the importance of the substitution at A ring, the long chain ester at the C8, the hydroxyl group at the C13, the phenyl ring at the C14 and the N-ethyl group, while the methoxy group at the C1 and C16 showed little effect on the activity. The results of the proliferation, apoptosis and ultrastructure tests of the treated canine mammary carcinoma cells referred that the representative compound, aconitine linoleate (25) could block the cell cycle of canine mammary carcinoma cells in the G0/G1 phase, and exhibit the anti-proliferative effect by inducing apoptosis.
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Alcaloides , Neoplasias de la Mama , Carcinoma , Diterpenos , Perros , Animales , Femenino , Humanos , Aconitina/farmacología , Aconitina/química , Neoplasias de la Mama/tratamiento farmacológico , Alcaloides/farmacología , Alcaloides/química , Diterpenos/farmacología , Diterpenos/químicaRESUMEN
P-glycoprotein (P-gp/ABCB1) and breast cancer resistance protein (BCRP/ABCG2) expression are frequently related to multidrug resistance (MDR) in neoplastic cells. Canine inflammatory and grade III noninflammatory mammary carcinomas (IMC and non-IMC) are aggressive tumors that could benefit from chemotherapy. This study describes the immunohistochemical detection of P-gp and BCRP in 20 IMCs and 18 non-IMCs from dogs that had not received chemotherapy. Our aim was to determine if P-gp and BCRP expression was related to the "inflammatory" phenotype, to establish a basis for future studies analyzing the response to chemotherapy in dogs with highly malignant mammary cancer. Immunolabeling was primarily membranous for P-gp with a more intense labeling in emboli, and immunolabeling was membranous and cytoplasmic for BCRP. P-gp was expressed in 17 of 20 (85%) IMCs compared to 7 of 18 (39%) non-IMCs (P = 0.006). BCRP was expressed within emboli in 15 of 19 (79%) emboli in IMC, 12 of 15 (80%) primary IMCs, and 12 of 18 (67%) non-IMCs, without statistically significant differences (P > .05). All IMCs and 67% of non-IMCs expressed at least 1 of the 2 transporters, and 63% (12/19) of IMCs and 39% (7/18) of non-IMCs expressed both P-gp and BCRP. P-gp and BCRP evaluation might help select patients for chemotherapy. P-gp, expressed in a significantly higher percentage of IMCs vs non-IMCs, might play a specific role in the chemoresistance of IMC.
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Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/metabolismo , Enfermedades de los Perros/patología , Neoplasias Mamarias Animales/patología , Animales , Enfermedades de los Perros/metabolismo , Perros , Femenino , Regulación Neoplásica de la Expresión Génica , Inmunohistoquímica/veterinaria , Neoplasias Mamarias Animales/metabolismo , Proteínas de Neoplasias/metabolismo , FenotipoRESUMEN
The presence of cancer stem-like cells (CSC) within canine mammary tumors, may explain partly local recurrence and spreading, since their ability to resist conventional antitumor treatments as chemo and radiotherapy. It has been recently described that simvastatin - a drug that inhibits synthesis of cholesterol - attenuates the proliferation of canine mammary CSC derived from CF41.Mg canine mammary carcinoma cells, promoting their chemosensitizing and apoptosis. The canonical Wnt/ß-catenin pathway is usually activated at CSC and up-regulates multidrug resistance protein 1 (MDR1), triggering chemoresistance. In the present study, we analyze the effect of simvastatin on ß-catenin/MDR1 expression in spheres obtained from the CF41.Mg cell line as a model of CSC. Simvastatin increased phosphorylation of ß-catenin without affecting its total expression. Moreover, MDR1 expression was decreased by simvastatin. These results suggest that simvastatin would facilitate the degradation of ß-catenin, decreasing MDR1 expression and contributing to the chemosensitizing effects of the statin on canine mammary CSC.
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Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Enfermedades de los Perros/metabolismo , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacología , Simvastatina/farmacología , Esferoides Celulares/efectos de los fármacos , beta Catenina/metabolismo , Animales , Línea Celular Tumoral , Perros , Femenino , Técnicas In Vitro , Neoplasias Mamarias Animales , Células Madre NeoplásicasRESUMEN
The CD44+/CD24- phenotype identifies cancer stem cell (CSC) properties in canine mammary carcinoma (MC); however, the histopathological features associated with this phenotype remain to be elucidated. Here, we determined whether the CD44+/CD24- phenotype was associated with hormonal receptor (HR; estrogen receptor [ER] and/or progesterone receptor [PR]) status and/or triple (ER, PR, and human epithelial growth factor receptor 2)-negative (TN) subtype; conventional histological evaluation was also performed. We found that, as single markers, both CD44+ and CD24+ were associated with less aggressive histological types, low grade, and a non-TN subtype; both markers were associated with HR positivity. On the other hand, a CD44+/CD24- phenotype was associated with higher grade of carcinoma. Therefore, our results suggest that immunohistochemical phenotyping for CD44/CD24 is useful for the evaluation of tumor behavior as well as CSC-like properties in canine MCs.
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Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/veterinaria , Neoplasias Mamarias Animales/patología , Células Madre Neoplásicas/metabolismo , Animales , Neoplasias de la Mama/patología , Antígeno CD24/metabolismo , Perros , Femenino , Receptores de Hialuranos/metabolismo , Inmunohistoquímica/veterinaria , Clasificación del Tumor , Receptores de Estrógenos/metabolismo , Receptores de Progesterona/metabolismoRESUMEN
Mammary tumour is the most common type of tumour in dogs, especially in unneutered female dogs. Homoharringtonine (HHT) is a natural alkaloid that can be used to treat various types of human tumour. However, the inhibitory effect and mechanism of HHT on canine mammary carcinomas (CMC) remain unclear. This study aimed to evaluate the inhibitory effect of HHT on CMC in vitro and determine its underlying molecular mechanism. The effects of HHT on the cytotoxicity of CMC U27 cells were evaluated by the cell counting kit-8, wound healing, and Transwell assays. HHT-induced apoptosis of U27 cells was detected by JC-1 and terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay. Moreover, the gene expression of B-cell lymphoma-2 (Bcl-2) and Bcl-2 associated X protein (Bax) were analysed using quantitative reverse transcription-polymerase chain reaction (RT-qPCR), and the protein expression of protein kinase B/mammalian target of rapamycin (AKT/mTOR) and mitochondrial apoptosis proteins were determined by western blotting. Furthermore, mammary tumour-bearing mouse models were established using 4T1 cells to evaluate the therapeutic effect of HHT. It was found that HHT could significantly down-regulated the protein expression of p-AKT, p-mTOR, and Bcl-2, and up-regulated the protein expression of P53, Bax, cleaved caspase-3, and cleaved caspase-9. In addition, HHT significantly suppressed both tumour volume and mass in mammary tumour mice. In conclusion, HHT damages CMC cells by inhibiting the AKT/mTOR signalling pathway and inducing mitochondrial apoptosis. Such findings lay a theoretical foundation for the clinical treatment of CMC and provide more options for clinical medication.
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Carcinoma , Enfermedades de los Perros , Enfermedades de los Roedores , Animales , Femenino , Perros , Humanos , Ratones , Homoharringtonina/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-akt/farmacología , Proteína X Asociada a bcl-2 , Enfermedades de los Perros/tratamiento farmacológico , Transducción de Señal , Apoptosis , Serina-Treonina Quinasas TOR/metabolismo , Carcinoma/veterinaria , Proliferación Celular , Mamíferos/metabolismoRESUMEN
Estrogen receptors (ERs) are located in both healthy and neoplastic tissues. The type of estrogen receptor expressed varies depending on its location, tumor type, and species. Estrogen action is mediated by binding to ER and activating the transcriptional and signaling processes that result in the control of gene expression. There are two main types of estrogen receptors: ER alpha (ERα) and ER beta (ERß). Both receptors are functionally different, they may act antagonistically and are distributed in different tissues but their structure is similar - as they are composed of 5 different domains: A/B, C, D, E, and F. The signaling pathway and hence regulation of the gene expression by ERs is a complex and multifactorial process that involves both genomic and nongenomic actions. In the human reproductive tract, both ERα and ß are present, with predominant expression of ERß, while there are no satisfactory data distinguishing the type of ERs expressed in the canine reproductive tract. In mammary gland neoplasia, a decreased or lacking ERα expression in humans is associated with a poorer prognosis. This is similar to dogs, where higher ERα expression intensity was noted in benign tumors than in carcinomas. In human hematopoietic malignancies, ERß is a predominant receptor. Selective and non-selective ERß agonists have an antiproliferative and pro-apoptotic effect on human lymphoma cell lines and may be effective in the therapy of ERß positive lymphomas and leukemias. In canine lymphoma tissues, none or only marginal expression of ERs was detected over the decades. Considering available data, we conducted preliminary studies proving that, in contrast to humans, the dominant ER expressed in canine hematopoietic tumors is ERα.
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Enfermedades de los Perros , Receptor alfa de Estrógeno , Receptor beta de Estrógeno , Neoplasias Hematológicas , Perros , Animales , Receptor beta de Estrógeno/genética , Receptor beta de Estrógeno/metabolismo , Receptor alfa de Estrógeno/genética , Receptor alfa de Estrógeno/metabolismo , Enfermedades de los Perros/genética , Enfermedades de los Perros/metabolismo , Neoplasias Hematológicas/veterinaria , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias/veterinaria , Neoplasias/genética , Neoplasias/metabolismoRESUMEN
Canine mammary carcinomas (CMC) are associated with major aggressive clinical behavior and high mortality. The current standard of care is based on surgical resection, without an established effective treatment scheme, highlighting the urgent need to develop novel effective therapies. Vascular endothelial growth factor (VEGF) is a key regulator of tumor angiogenesis and progression in the majority of solid cancers, including human and canine mammary carcinomas. The first therapy developed to target VEGF was bevacizumab, a recombinant humanized monoclonal antibody, which has already been approved as an anticancer agent in several human cancers. The goal of this work was to establish the therapeutic value of MB02 bevacizumab biosimilar in CMC. First, through different in silico approaches using the MUSCLE multiple-sequence alignment tool and the FoldX protein design algorithm, we were able to predict that canine VEGF is recognized by bevacizumab, after showing an extremely high sequence similarity between canine and human VEGF. Further, by using an ELISA-based in vitro binding assay, we confirmed that MB02 biosimilar was able to recognize canine VEGF. Additionally, canine VEGF-induced microvascular endothelial cell proliferation was inhibited in a concentration-dependent manner by MB02 biosimilar. These encouraging results show a high potential for MB02 as a promising therapeutic agent for the management of CMC.
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Spontaneous canine mammary carcinomas (CMCs) have been widely considered a good research model for human breast cancers, which brings much attention to CMCs. In recent years, the oncolytic effect of Newcastle disease virus (NDV) on cancer cells has been widely studied, but its effect on CMCs is still unclear. This study aims to investigate the oncolytic effect of NDV LaSota strain on canine mammary carcinoma cell line (CMT-U27) in vivo and in vitro. The in vitro cytotoxicity and immunocytochemistry experiments showed that NDV selectively replicated in CMT-U27 cells, and inhibited cell proliferation and migration but not in MDCK cells. KEGG analysis of transcriptome sequencing indicated the importance of the TNFα and NF-κB signalling pathways in the anti-tumour effect of NDV. Subsequently, the significantly increased expression of TNFα, p65, phospho-p65, caspase-8, caspase-3 and cleaved-PARP proteins in the NDV group suggested that NDV induced CMT-U27 cells apoptosis by activating the caspase-8/caspase-3 pathway and the TNFα/NF-κB signalling pathway. Nude mice tumour-bearing experiments showed that NDV could significantly decrease the growth rate of CMC in vivo. In conclusion, our study demonstrates the effective oncolytic effects of NDV on CMT-U27 cells in vivo and in vitro, and suggests NDV as a promising candidate for oncolytic therapy.
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Carcinoma , Enfermedades de los Perros , Viroterapia Oncolítica , Virus Oncolíticos , Animales , Perros , Ratones , Apoptosis , Carcinoma/veterinaria , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Línea Celular Tumoral , Enfermedades de los Perros/terapia , Ratones Desnudos , Virus de la Enfermedad de Newcastle/metabolismo , FN-kappa B/metabolismo , Viroterapia Oncolítica/veterinaria , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Introduction: Enhancer of zeste homologue 2 (EZH2) is the human homologue of Drosophila zeste gene enhancer. The aim of this study was to determine the expression of EZH2 in canine mammary carcinomas (CMCs) and its relationship with clinicopathological features. Material and Methods: The expression of EZH2 mRNA and protein in 53 CMC tissue and 8 normal mammary gland tissue samples was measured by quantitative real-time PCR and immunohistochemical staining assay, respectively. The relationship between EZH2 protein expression and clinicopathological features was analysed by χ2 test to further explore the clinical significance of EZH2 in CMCs. Results: Compared with normal mammary gland tissues, EZH2 mRNA expressions were significantly increased in CMC tissues (P < 0.01). Moreover, normal mammary glands did not express the EZH2 protein but carcinomic glands did, and expression increased in CMCs with high histological grades, especially in histological grade II (P < 0.05). However, EZH2 expression was not related to age, tumour size, or metastasis (P > 0.05). The expression of EZH2 in one type of CMC was not significantly different from the expression in any other type (P > 0.05). Conclusion: EZH2 is highly expressed in CMCs, indicating that it can be used as a molecular marker for early diagnosis, prognosis, or therapy of CMCs.
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Neoplastic cells, through immunoediting mechanisms, can establish a state of immunosuppression to evade host immune defenses. The aims of this study were: (1) to validate a standard method for assessing tumor infiltrating lymphocytes (TILs) in canine mammary carcinoma by applying international human breast cancer guidelines; (2) to investigate if the TILs population was composed of a subset of regulatory T lymphocytes (Tregs); and (3) to evaluate the relationship between the number of TILs and Tregs and the biological behavior of the tumors. One hundred and twenty-nine canine mammary tumors were retrospectively selected for this study. Histological diagnosis, grading and histological evaluation of TILs was performed on hematoxylin and eosin-stained sections. TILs were evaluated using a three-tier semiquantitative method, previously validated in human medicine, based on the percentage of TILs (0-10%, 11-40% and 41-90%). Lymphocyte immunophenotype was confirmed by CD3 and CD79, while an anti-FoxP3 antibody was used to determine the presence of Tregs. The number of stromal TILs and invasive front TILs significantly correlated with each other (P < 0.0001) and increased with increasing histological grade (P = 0.002 and P = 0.004, respectively). A subset of TILs was composed of FOXP3+ Tregs. Stromal Tregs and invasive front Tregs were associated with stromal TILs and invasive front TILs (P = 0.03; P = 0.01 and P = 0.003; P = 0.007, respectively). In conclusion, in canine mammary carcinomas, an increased number of stromal and invasive front TILs is associated with increased malignancy and significant increase of Tregs that could lead to immunosuppression and evasion of the host immune system.
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Neoplasias de la Mama , Carcinoma , Enfermedades de los Perros , Animales , Neoplasias de la Mama/veterinaria , Carcinoma/patología , Carcinoma/veterinaria , Enfermedades de los Perros/patología , Perros , Femenino , Inmunofenotipificación/veterinaria , Linfocitos Infiltrantes de Tumor/patología , Pronóstico , Estudios Retrospectivos , Linfocitos T ReguladoresRESUMEN
Human epidermal growth factor receptor 2 (HER2) is a tyrosine kinase receptor that promotes tumor cell growth and is implicated in the pathogenesis of human breast cancer. The role of HER2 in canine mammary carcinomas (CMCs) is not clear. Therefore, this study aimed to examine the protein expression and cytogenetic changes of HER2 and their correlation with other clinical-pathological parameters in CMC. We retrospectively selected 112 CMCs. HER2, ER, and Ki67 were assessed by immunohistochemistry. HER2 antibody validation was investigated by immunoblot on mammary tumor cell lines. Fluorescence in situ hybridization (FISH) was performed with probes for HER2 and CRYBA1 (control gene present on CFA9). HER2 protein overexpression was detected in 15 carcinomas (13.5%). A total of 90 carcinomas were considered technically adequate by FISH, and 8 out of 90 CMC (10%) were HER2 amplified, 3 of which showed a cluster-type pattern. HER2 overexpression was correlated with an increased number of HER2 gene copies (p = 0.01; R = 0.24) and overall survival (p = 0.03), but no correlation with ER, Ki67, grade, metastases, and tumor-specific survival was found. Surprisingly, co-amplification or polysomy was identified in three tumors, characterized by an increased copy number of both HER2 and CRYBA1. A morphological translocation-fusion pattern was recognized in 20 carcinomas (22%), with a co-localized signal of HER2 and CRYBA1. HER2 is not associated with clinical-pathological parameters of increased malignancy in canine mammary tumors, but it is suitable for studying different amplification patterns.
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Canine mammary carcinoma (CMC) is the most common type of neoplasm in intact female dogs. While a previous study in Western countries validated the 2011 classification as an independent prognostic indicator in CMC, its role in CMC prognostication in Asian countries such as Korea remains unclear. In the present study, we estimate the survival rates in CMC types defined by the 2011 classification, elucidate the prognostic significance of the histological subtype and grade and that of the lymphatic invasion status in CMC, and validate the 2011 classification as an independent prognostic indicator in a large cohort of CMCs (excluding cases of multicentric CMCs). A total of 155 CMC cases retrieved from archived formalin-fixed, paraffin-embedded tissues, along with 2-year follow-up data, were retrospectively analysed. A significant association was found between the histological subtype of the 2011 classification and the tumour-specific survival. Carcinosarcoma, adenosquamous carcinoma and anaplastic carcinoma subtypes were associated with the poorest prognosis. Dogs with comedocarcinoma and solid carcinoma followed a disease course that was more aggressive than that observed in dogs with a carcinoma arising in a benign mixed tumour. Moreover, age, histological grade and lymphatic invasion status significantly correlated with tumour-specific survival in univariate analysis. In multivariate analysis, histological subtype, age and lymphatic invasion status remained independent prognostic factors for CMC.
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Carcinoma , Enfermedades de los Perros , Neoplasias Mamarias Animales , Animales , Carcinoma/patología , Carcinoma/veterinaria , Enfermedades de los Perros/patología , Perros , Femenino , Neoplasias Mamarias Animales/patología , Pronóstico , Estudios RetrospectivosRESUMEN
Canine mammary gland tumors (CMTs) are one of the most prevalent cancers in dogs and a good model for human breast cancer (BC), however gene expression analysis of CMTs is scarce. Although divergence of genes expression has been found in BC of different human races, no such research of different dog's breeds has been done. The purpose of this study was to investigate expression of the VEGF-B, VEGF-A, FLT-1, KDR, ERBB2, EGFR, GRB2, RAC1, CDH1 and HYAL-1 genes of canine mammary carcinomas, compare the expression levels with clinicopathological parameters and analyze expression disparities between different breeds. Carcinomas and adjacent tissues were collected from female dogs to perform routine histopathology, immunohistochemistry (IHC) and quantitative real-time polymerase chain reaction (qRT-PCR). We found that VEGF-B and EGFR genes were overexpressed in the mammary gland carcinomas compared to adjacent tissue. VEGF-B gene expression had associations with different parameters (tumor size, grade, and absence of metastasis). Furthermore, differences in VEGF-B, FLT1, ERBB2, GRB2, RAC1, CDH1 and HYAL-1 genes expression have been found in different breed dogs (German Shepherd, Yorkshire Terrier) and mixed-breed dogs indicating that a dog's breed could determine a molecular difference, outcome of cancer and should be accounted as a confounding factor in the future gene expression research.
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P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) are major actors in multidrug resistance (MDR) phenomenon in both human and canine mammary carcinomas (CMCs). The aim of this study was to investigate an association between the intrinsic expression of P-gp and BCRP compared to the immunophenotypes and outcome in CMCs. Fifty CMCs were evaluated at immunohistochemistry (IHC) for P-gp, BCRP, Estrogen receptor alpha (ER), Progesterone receptors (PR), Human Epidermal Growth Factor Receptor type 2 (HER2), basal cytokeratins 5/6 (CK5/6), Epidermal Growth Factor Receptor 1 (EGFR), and Ki67 proliferation index. P-gp and BCRP positive cases were, respectively, 52% and 74.5%, with a significantly higher expression of BCRP than P-gp. Five immunophenotypes were defined in 37 out of 50 CMCs: 9 (24.3%) Luminal A, 5 (13.5%) Luminal B, 9 (24.3%) HER2 overexpressing, 9 (24.3%) Triple-negative basal-like, and 5 (13.5%) Triple-negative non-basal-like. In all CMCs at least one marker was expressed. Follow-up data were available for 25 animals. The average cancer-specific survival was 739 ± 444 days. A number of CMCs bear a high expression of P-gp and BCRP but no significant association was found between their expression and the immunophenotypes, Ki67 index, the histological grade, and tumor-related death.
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Cancer-associated stroma (CAS) profoundly influences progression of tumors including mammary carcinoma (mCA). Canine simple mCA represent relevant models of human mCA, notably also with respect to CAS. While transcriptomic changes in CAS of mCA are well described, it remains unclear to what extent these translate to the protein level. Therefore, we sought to gain insight into the proteomic changes in CAS and compare them with transcriptomic changes in the same tissue. To this end, we analyzed CAS and matched normal stroma using laser-capture microdissection (LCM) and LC-MS/MS in a cohort of 14 formalin-fixed paraffin embedded (FFPE) canine mCAs that we had previously characterized using LCM-RNAseq. Our results reveal clear differences in protein abundance between CAS and normal stroma, which are characterized by changes in the extracellular matrix, the cytoskeleton, and cytokines such as TNF. The proteomics- and RNAseq-based analyses of LCM-FFPE show a substantial degree of correlation, especially for the most deregulated targets and a comparable activation of pathways. Finally, we validate transcriptomic upregulation of LTBP2, IGFBP2, COL6A5, POSTN, FN1, COL4A1, COL12A1, PLOD2, COL4A2, and IGFBP7 in CAS on the protein level and demonstrate their adverse prognostic value for human breast cancer. Given the relevance of canine mCA as a model for the human disease, our analysis substantiates these targets as disease-promoting stromal components with implications for breast cancer in both species.
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Fibroblastos Asociados al Cáncer/patología , Neoplasias Mamarias Animales/patología , Células del Estroma/patología , Transcriptoma/genética , Animales , Citocinas/metabolismo , Citoesqueleto/fisiología , Modelos Animales de Enfermedad , Perros , Matriz Extracelular/fisiología , Femenino , Perfilación de la Expresión Génica , Captura por Microdisección con Láser , Espectrometría de Masas en Tándem , Microambiente Tumoral/fisiología , Regulación hacia ArribaRESUMEN
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.
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Biomarcadores de Tumor/metabolismo , Carcinoma/veterinaria , Enfermedades de los Perros/metabolismo , Neoplasias Mamarias Animales/metabolismo , Animales , Carcinoma/metabolismo , Carcinoma/mortalidad , Carcinoma/patología , Ciclooxigenasa 2/metabolismo , Enfermedades de los Perros/mortalidad , Enfermedades de los Perros/patología , Perros , Receptores ErbB/metabolismo , Femenino , Inmunohistoquímica/veterinaria , Neoplasias Mamarias Animales/mortalidad , Metástasis de la Neoplasia , Estadificación de Neoplasias/veterinaria , Pronóstico , Estudios Prospectivos , Receptores de Estrógenos/metabolismo , Receptores de Progesterona/metabolismo , EspañaRESUMEN
Canine mammary carcinomas are common tumours in female dogs and histopathological examination has an important role in identifying whether they are benign or malignant. The latest and most commonly used histological grading system was established by Peña et al. (2013) and is based on the extent of tubule formation, nuclear pleomorphism and number of mitoses. Before the establishment of this grading system, tumour size and classical histological indicators of malignancy such as lymphovascular invasion, infiltration into surrounding tissue, necrosis and presence of a micropapillary pattern were important predictors of biological behaviour. However, the system of Peña et al. does not consider tumour size or these histological features. Clarifying the association of these features and histological grade, especially in grade II and III carcinomas, is important. In this study, we confirmed that the system of Peña et al. is effective for predicting biological behaviour and that evaluation of histological features of malignancy reinforced histological grade, as determined by the system of Peña et al., especially in grade II carcinomas.
Asunto(s)
Enfermedades de los Perros/patología , Neoplasias Mamarias Animales/patología , Animales , Perros , Femenino , Clasificación del TumorRESUMEN
By using the Warburg effect-a phenomenon where tumors consume higher glucose levels than normal cells-on cancer cells to enhance the effect of photodynamic therapy (PDT), we developed a new photosensitizer, glucose-conjugated chlorin e6 (G-Ce6). We analyzed the efficacy of PDT with G-Ce6 against canine mammary carcinoma (CMC) in vitro and in vivo. The pharmacokinetics of G-Ce6 at 2, 5, and 20 mg/kg was examined in normal dogs, whereas its intracellular localization, concentration, and photodynamic effects were investigated in vitro using CMC cells (SNP cells). G-Ce6 (10 mg/kg) was administered in vivo at 5 min or 3 h before laser irradiation to SNP tumor-bearing murine models. The in vitro study revealed that G-Ce6 was mainly localized to the lysosomes. Cell viability decreased in a G-Ce6 concentration- and light intensity-dependent manner in the PDT group. Cell death induced by PDT with G-Ce6 was not inhibited by an apoptosis inhibitor. In the in vivo study, 5-min-interval PDT exhibited greater effects than 3-h-interval PDT. The mean maximum blood concentration and half-life of G-Ce6 (2 mg/kg) were 15.19 ± 4.44 µg/mL and 3.02 ± 0.58 h, respectively. Thus, 5-min-interval PDT with G-Ce6 was considered effective against CMC.