T Lymphocytes in Histiocytic Sarcomas of Flat-Coated Retriever Dogs.

Flat-Coated Retriever dogs are predisposed to the development of histiocytic sarcoma (HS), a poorly differentiated, highly malignant neoplasm. The authors have previously documented a significant lymphocytic infiltrate in such tumors. The objective of this study was to examine the presence and expression of regulatory T cells in HS tumor samples. Forty tumors were included in this study. All tumors were immunolabeled for CD3, CD79a, CD25, CD45RA, and FOXP3. The proportion of positive cells was compared between tumors presenting as a localized primary soft tissue mass (soft tissue origin HS) and disseminated HS affecting viscera, especially the spleen (splenic origin HS). By immunohistochemistry, 95% of infiltrating T cells were positive for Foxp3 in all sections, suggesting the presence of regulatory T cells. The proportion of cells positive for FOXP3 was higher in the tumors arising in soft tissues, whereas the proportion of CD45RA-positive cells was higher in the splenic origin HS. Canine HS has an aggressive clinical behavior and is uniformly fatal. The difference in the proportion of tumor-infiltrating lymphocytes positive for these 2 markers in the 2 locations may represent differences in tumor microenvironment between the 2 sites.

Expression of Fibroblast Activating Protein and Correlation with Histological Grade, Mitotic Index and Ki67 Expression in Canine Mast Cell Tumours.

Fibroblast activating protein (FAP) is a membrane serine protease expressed by activated fibroblasts, particularly tumour associated fibroblasts (TAFs). FAP expression has not been reported in canine mast cell tumours (MCTs). The objective of this study was to evaluate the expression of FAP in TAFs and its correlation with histological grade, mitotic index and Ki67 expression in canine MCTs. FAP expression was evaluated by immunohistochemistry (IHC) in 30 canine MCTs. Twenty-eight (90%) of the MCTs expressed FAP in the stroma, 16 cases showed low to intermediate FAP score and 14 cases had a high FAP score. FAP was correlated positively with both Patnaik (P = 0.007) and Kiupel (P = 0.008) grading systems, mitotic index (P = 0.0008) and Ki67 expression (P = 0.009). High stromal FAP expression could be a potential negative prognostic factor in canine MCTs.

An immunohistochemical study of the expression of the hypoxia markers Glut-1 and Ca-IX in canine sarcomas.

Tumor hypoxia has been associated with increased malignancy, likelihood of metastasis, and increased resistance to radiotherapy and chemotherapy in human medicine. Hypoxia-inducible factor-1 (HIF-1) is a key transcription factor that is induced by tumor hypoxia and regulates the pathways involved in cellular response and adaptation to the hostile tumor microenvironment. HIF-1 induces transcription of different proteins, including Ca-IX and Glut-1, which are considered endogenous markers of chronic hypoxia in solid tumors in humans. In this study, sections from 40 canine sarcomas (20 histiocytic sarcomas and 20 low-grade soft-tissue sarcomas) were immunostained for these markers. Expression of Glut-1 was scored based on percentage of positive staining cells (0 = <1%; 1 = 1%-50%; 2 = >50%) and intensity of cellular staining (1 = weak; 2 = strong); Ca-IX was scored based on percentage of positive cells (0 = <1%; 1 = 1%-30%; 2 = >30%). Intratumoral microvessel density was measured using CD31 to assess intratumoral neoangiogenesis. Histiocytic sarcomas showed statistically significant higher Glut-1 immunoreactivity and angiogenesis than did low-grade soft-tissue sarcomas. Intratumoral microvessel density in histiocytic sarcomas was positively associated with Glut-1 immunoreactivity score. These findings suggest a potential role of hypoxia in the biology of these tumors and may provide a base for investigation of the potential prognostic use of these markers in naturally occurring canine tumors.

The clinical presentation and histopathologic-immunohistochemical classification of histiocytic sarcomas in the Flat Coated Retriever.

The Flat Coated Retriever is a breed at risk of development of histiocytic sarcoma (HS), but in contrast to the disseminated form of disease recognized in the Bernese Mountain Dog, most reports of HS in Flat Coated Retrievers describe a localized lesion affecting the musculature or fascia of limbs. The purpose of this study was to review data and material received though an ongoing Flat Coated Retriever tumor survey to better define the presentation of HS in the breed and to determine the utility of subclassification of tumors arising at different sites by histology and immunohistologic phenotyping. Data on 180 dogs bearing HS-like tumors were available for review, which showed that although the majority (101 lesions, 57%) were primary limb lesions, 47 dogs (26%) had visceral, mainly splenic lesions with no peripheral primary tumor. A detailed histologic and immunohistologic review of 20 limb tumors and 20 splenic tumors showed that 2 distinct phenotypic subtypes could be identified: a histiocytic subtype, most prevalent in the splenic tumors, and a histiocytic-spindle-pleomorphic subtype, mainly seen in the limb tumors. Despite their variable morphology, all tumors expressed major histocompatibility complex class II and the leukocyte antigen CD18, but only those tumors in the spleen consistently expressed CD11d. The majority of tumors also contained a mild to moderate infiltrate of T lymphocytes.

TSLC1 tumour-suppressor gene expression in canine mast cell tumours.

Tumour suppressor in lung cancer-1 (TSLC1) is a tumour-suppressor gene coding for an adhesion molecule that is expressed by mast cells. Reduced TSLC1 expression is associated with a poor prognosis in several human tumours, and this study sought to investigate if TSLC1 expression could be used to predict outcome in dogs with mast cell tumours (MCTs). Sections of MCTs of different tumour grades from 45 dogs (Group 1) were immunohistochemically assessed for TSLC1 and Ki67 expression. In addition, 35 intermediate-grade MCTs (Group 2) from dogs with known clinical follow-up were immunohistochemically stained for TSLC1 and Ki67. The TSLC1 staining intensity was found to strongly inversely correlate with tumour grade for Group 1 (P = 0.002857). For Group 2 there was a trend towards dogs with lower TSLC1 scores being more likely to die from MCT-related disease (P = 0.058). The intensity of TSLC1 staining inversely correlated with Ki67 expression for both groups.

Chlorambucil and prednisolone chemotherapy for dogs with inoperable mast cell tumours: 21 cases.

OBJECTIVES: To evaluate the response of measurable canine mast cell tumours unsuitable for other treatment modalities to a chemotherapy protocol comprising chlorambucil and prednisolone. METHODS: Dogs bearing measurable mast cell tumours, unsuitable for treatment by surgery or radiotherapy, were treated with orally administered prednisolone and chlorambucil, and their responses assessed. RESULTS: Twenty-one dogs were enrolled in the study; 13 had intermediate-grade mast cell tumour, six were high grade and two were diagnosed by cytology alone. Eight dogs had multiple tumours and 13 dogs had single tumours, and six dogs had lymph node metastases and no dogs had visceral metastases detected. Three dogs achieved complete remission, five achieved partial remission (overall response rate 38 per cent), nine had static disease and four dogs had progressive disease. Median progression-free interval for the eight responders was 533 days, and median survival time for all dogs in the study was 140 days. Progression-free interval and median survival time were not influenced by the age, sex, weight or neutering status of the patient, by the grade or stage of the tumour or whether the patient had single or multiple tumours. No toxicity was detected. CLINICAL SIGNIFICANCE: Response and survival rates of inoperable canine MCT to chlorambucil and prednisolone are comparable to previously described protocols, with no apparent toxicity.

Mortality in a cohort of flat-coated retrievers in the UK.

A cohort study of 174 flat-coated retrievers was undertaken to establish the importance of cancer in flat coat mortality in terms of the prevalence of neoplasia in the breed and also the relative effect of cancer on lifespan in relation to other forms of mortality. Dogs aged 2-7 years were recruited in 1996 and followed until 2007. An annual health census was used to collect the data. Two dogs were lost to follow-up and 72 dogs (42%) died from confirmed neoplasia. Twenty dogs (11.6%) died of unconfirmed tumours and 61 (35%) died from non-neoplastic conditions. The cause of death was unknown for 19 dogs. Soft tissue sarcoma (especially histiocytic sarcoma) was the predominant cancer type, affecting 32 dogs (44% of neoplasms). Six dogs died with malignant melanoma and three with lymphoma. Median age at death was 9 years for dogs with tumours (eight for sarcoma patients) and 12 years for non-neoplastic fatalities. The results confirm that soft tissue sarcoma, particularly histiocytic sarcoma, is a major cause of mortality in this breed.

Histiocytic sarcoma of the spleen in flat-coated retrievers with regenerative anaemia and hypoproteinaemia.

Three flat-coated retrievers with a regenerative anaemia were examined. They were hypoproteinaemic suggesting that the anaemia might be due to blood loss, but it was not possible to identify a site of haemorrhage. All three had splenomegaly with splenic abnormalities apparent on ultrasonography. Ultimately all three animals were shown to have a histiocytic sarcoma involving the spleen and other tissues. A fourth flat-coated retriever with anaemia, hypoproteinaemia and an abdominal mass was also diagnosed with a histiocytic sarcoma of the spleen following splenectomy. It is postulated that the dogs' anaemia was due to erythrophagocytosis, either directly by neoplastic cells or by reactive macrophages.

Chromosome rearrangements in canine fibrosarcomas.

We have previously reported the use of six- and seven-color paint sets in the analysis of canine soft tissue sarcomas. Here we combine this technique with flow sorting of translocation chromosomes, reverse painting, and polymerase chain reaction (PCR) analysis of the gene content of the reverse paint in order to provide a more detailed analysis of cytogenetic abnormalities in canine tumors. We examine two fibrosarcomas, both from female Labrador retrievers, and show abnormalities in chromosomes 11 and 30 in both cases. Evidence of involvement of TGFBR1 is presented for one tumor.

Immunohistochemical and histopathologic features of 14 malignant fibrous histiocytomas from Flat-Coated Retrievers.

Flat-Coated Retrievers seem to be at increased risk of developing soft-tissue sarcomas, and undifferentiated round cell or spindle cell sarcomas account for approximately 59% of sarcomas in the breed. In an attempt to classify these tumors further, formalin-fixed, paraffin-embedded sections from 14 undifferentiated sarcomas from Flat-Coated Retrievers were reviewed and examined with a panel of histologic and immunohistochemical stains. The panel included vimentin, desmin, Myo D1, smooth muscle actin, cytokeratin, S100, von Willebrand factor (factor VIII), Mac 387, CD3, major histocompatibility complex (MHC) class II, and CD79a. The majority of the sarcomas showed greater than 70% staining for MHC class II. We conclude that these undifferentiated sarcomas in Flat-Coated Retrievers belong to a spectrum of tumors with varying proportions of characteristic cell types and morphologic features, some of which fit the diagnostic criteria for malignant fibrous histiocytoma. Many of these sarcomas seem to have a significant myofibroblast component and a mild or moderate T cell infiltrate but the precise cell lineage is still uncertain.

Prognostic variables in canine multicentric lymphosarcoma.

This paper presents the results of a prospective study to investigate the prognostic value of clinical staging, histological grading, immunophenotype, mitotic count and average numbers of argyrophilic nucleolar organiser region counts in dogs with multicentric lymphosarcoma treated with a standard chemotherapy protocol comprising vincristine, cyclophosphamide and prednisolone. Forty-nine dogs were treated according to the study protocol. Univariate and multivariate analysis with regression modelling was used to evaluate the prognostic importance of patient and tumour variables upon tumour response and relapse-free survival. Thirty-seven dogs (76 per cent) achieved a complete remission, seven (14 per cent) a partial remission and five (10 per cent) failed to respond to treatment. None of the variables examined had a statistically significant effect upon tumour response. Tumour immunophenotype was the only parameter found to have a significant influence on patient survival, the hazard ratio for T-cell versus B-cell immunophenotype was 3.99 with 95 per cent confidence interval from 1.399 to 11.372, P = 0.035.

Histopathological survey of neoplasms in flat-coated retrievers, 1990 to 1998.

Over the period from March 1990 to December 1998, veterinary surgeons in general practice were invited to submit tissues suspected of being neoplastic which had been removed from flat-coated retrievers. When possible, pedigree details were obtained from the owners. In addition, data were collected from flat-coated retrievers known to have suffered from a neoplastic condition and for which a histopathological report was available. A total of 1023 submissions was obtained from 782 dogs. These included 165 non-neoplastic lesions (16 per cent), 447 benign samples (44 per cent) and 411 malignant samples (40 per cent). Soft tissue sarcomas accounted for 55 per cent of the malignant samples (26 per cent of all tumour samples and 22 per cent of all submissions) with 63 per cent of them being diagnosed as undifferentiated. Carcinomas accounted for 20 per cent of malignant samples (8 per cent of all submissions). Of the benign tumours, cutaneous histiocytoma was the most common diagnosis (48 per cent of benign tumours, 25 per cent of all tumours and 21 per cent of all submissions).