Currently, prognostic and therapeutic determinations for canine cutaneous mast cell tumors (MCTs) are primarily based on histologic grade. However, the use of different grading systems by veterinary pathologists and institutional modifications make the prognostic value of histologic grading highly questionable. To evaluate the consistency of microscopic grading among veterinary pathologists and the prognostic significance of the Patnaik grading system, 95 cutaneous MCTs from 95 dogs were

A study was carried out to test the accuracy and consistency of veterinary pathologists, not specialists in hematopathology, in applying the World Health Organization (WHO) system of classification of canine lymphomas. This study represents an initiative of the ACVP Oncology Committee, and the classification has been endorsed by the World Small Animal Veterinary Association (WASVA). Tissue biopsies from cases of canine lymphoma were received from veterinary oncologists, and a study by pathologists given only signalment was carried out on 300 cases. Twenty pathologists reviewed these 300 cases with each required to choose a diagnosis from a list of 43 B and T cell lymphomas. Three of the 20 were hematopathologists who determined the consensus diagnosis for each case. The 17 who formed the test group were experienced but not specialists in hematopathology, and most were diplomates of the American or European Colleges of Veterinary Pathology. The overall accuracy of the 17 pathologists on the 300 cases was 83%. When the analysis was limited to the 6 most common diagnoses, containing 80% of all cases, accuracy rose to 87%. In a test of reproducibility enabled by reintroducing 5% of cases entered under a different identity, the overall agreement between the first and second diagnosis ranged from 40 to 87%. The statistical review included 43,000 data points for each of the 20 pathologists.

Abstract. Canine cutaneous mast cell tumor (MCT) is a common neoplastic disease in dogs. Due to the prevalence of canine MCTs and the variable biologic behavior of this disease, accurate prognostication and a thorough understanding of MCT biology are critical for the treatment of this disease. The goals of this study were to evaluate and compare the utility of the proliferation markers Ki67, proliferating cell nuclear antigen (PCNA), and argyrophilic nucleolar organizing region (AgNOR) as independent prognostic markers for canine MCTs and to evaluate the use of these markers in combination, as each marker assesses different aspects of cellular proliferation. An additional goal of this study was to evaluate the associations between cellular proliferation and c-KIT mutations and between cellular proliferation and aberrant KIT protein localization in canine MCTs. Fifty-six MCTs treated with surgical excision alone were included in this study. Each MCT was evaluated for Ki67 expression, PCNA expression, and KIT protein localization using immunohistochemistry; for AgNOR counts using histochemical staining; and for the presence of internal tandem duplication c-KIT mutations using polymerase chain reaction amplification. In this study, increased Ki67 and AgNOR counts were both associated with significantly decreased survival. On the basis of these results, we recommend that the evaluation of cellular proliferation, including evaluations of both Ki67 expression and AgNORs, should be routinely used in the prognostication of canine MCTs. Additionally, the results of this study show that MCTs with aberrant KIT protein localization or internal tandem duplication c-KIT mutations are associated with increased cellular proliferation, further suggesting a role for c-KIT in the progression of canine MCTs.

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The c-KIT proto-oncogene has been implicated in the pathogenesis of several neoplastic diseases, including gastrointestinal stromal tumors and mastocytosis in humans, and mast cell tumors (MCTs) in canines. Cutaneous MCTs are common neoplasms in dogs and have a variable biologic behavior. The goal of this study was to define the prognostic significance of c-KIT mutations identified in canine MCTs and the associations between c-KIT mutations, KIT localization, and KIT expression levels. Microdissection and polymerase chain reaction were performed on 60 MCTs to identify c-KIT mutations. Anti-KIT antibodies were used for immunohistochemical evaluation of KIT localization. Forty-two MCTs were included in a tissue microarray, and KIT expression was quantified using immunofluorescence. Canine MCTs with c-KIT mutations were significantly associated with an increased incidence of recurrent disease and death. c-KIT mutations were also significantly associated with aberrant protein localization; however, the level of KIT expression did not correlate with either c-KIT mutations or changes in protein localization. Considering the high prevalence of canine MCTs and the central role of c-KIT in the tumorigenesis of certain tumors, canine MCTs are an excellent model for characterizing the role of c-KIT in neoplastic diseases and is a potential target for novel therapeutic agents in clinical trials.

Abstract. This document is the consensus of the American Association of Veterinary Laboratory Diagnosticians (AAVLD) Subcommittee on Standardization of Immunohistochemistry on a set of guidelines for immunohistochemistry (IHC) testing in veterinary laboratories. Immunohistochemistry is a powerful ancillary methodology frequently used in many veterinary laboratories for both diagnostic and research purposes. However, neither standardization nor validation of IHC tests has been completely achieved in veterinary medicine. This document addresses both issues. Topics covered include antibody selection, fixation, antigen retrieval, antibody incubation, antibody dilutions, tissue and reagent controls, buffers, and detection systems. The validation of an IHC test is addressed for both infectious diseases and neoplastic processes. In addition, storage and handling of IHC reagents, interpretation, quality control and assurance, and troubleshooting are also discussed. Proper standardization and validation of IHC will improve the quality of diagnostics in veterinary laboratories.

Abstract. Cutaneous mast cell tumors (MCTs) are one of the most common tumors in dogs. Currently, prognostic and therapeutic determinations for MCTs are primarily based on the histologic grade of the tumor, but a vast majority of MCTs are of an intermediate grade, and the prognostic relevance is highly questioned. A more detailed prognostic evaluation, especially of grade 2 canine MCTs, is greatly needed. To evaluate the prognostic significance of KIT and tryptase expression patterns in canine cutaneous MCTs, we studied 100 cutaneous MCTs from 100 dogs that had been treated with surgery only. The total survival and disease-free survival time and the time to local or distant recurrence of MCTs were recorded for all dogs. Using immunohistochemistry, 98 of these MCTs were stained with anti-KIT and antitryptase antibodies. Three KIT-and three tryptase-staining patterns were identified. The KIT-staining patterns were identified as 1) membrane-associated staining, 2) focal to stippled cytoplasmic staining with decreased membrane-associated staining, and 3) diffuse cytoplasmic staining. The tryptase-staining patterns were identified as 1) diffuse cytoplasmic staining, 2) stippled cytoplasmic staining, and 3) little to no cytoplasmic staining. Based on univariate and multivariate survival analysis, increased cytoplasmic KIT staining was significantly associated with an increased rate of local recurrence and a decreased survival rate. The tryptase-staining patterns were not significantly associated with any survival parameter. On the basis of these results, we propose a new prognostic classification of canine cutaneous MCTs, according to their KIT-staining pattern, that can be used for the routine prognostic evaluation of canine cutaneous MCTs.