Stereology in Grading and Prognosis of Canine Cutaneous Mast Cell Tumors.
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ABSTRACT:
SUBMITTER: Casanova M
PROVIDER: S-EPMC8064533 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Mutations in the c-KIT proto-oncogene have been implicated in the progression of several neoplastic diseases, including gastrointestinal stromal tumors and mastocytosis in humans, and cutaneous mast cell tumors (MCTs) in canines. Mutations in human mastocytosis patients primarily occur in c-KIT exon 17, which encodes a portion of its kinase domain. In contrast, deletions and internal tandem duplication (ITD) mutations are found in the juxtamembrane domain of c-KIT in approximately 15% of canine MCTs. In addition, ITD c-KIT mutations are significantly associated with aberrant KIT protein localization in canine MCTs. However, some canine MCTs have aberrant KIT localization but lack ITD c-KIT mutations, suggesting that other mutations or other factors may be responsible for aberrant KIT localization in these tumors.In order to characterize the prevalence of mutations in the phospho-transferase portion of c-KIT's kinase domain in canine MCTs exons 16-20 of 33 canine MCTs from 33 dogs were amplified and sequenced. Additionally, in order to determine if mutations in c-KIT exon 17 are responsible for aberrant KIT localization in MCTs that lack juxtamembrane domain c-KIT mutations, c-KIT exon 17 was amplified and sequenced from 18 canine MCTs that showed an aberrant KIT localization pattern but did not have ITD c-KIT mutations.No mutations or polymorphisms were identified in exons 16-20 of any of the MCTs examined.In conclusion, mutations in the phospho-transferase portion of c-KIT's kinase domain do not play an important role in the progression of canine cutaneous MCTs, or in the aberrant localization of KIT in canine MCTs.
| S-EPMC1448201 | biostudies-literature
Project description:Mast cell tumors (MCTs) are the most common skin tumor of the dog, and accurately predicting their clinical behavior is critical in directing patient therapy, as they range from benign lesions to a fatal systemic disease. Grading is useful for prognosis, but it cannot predict the behavior of all MCTs. We hypothesized that biomarker immunolabeling in tumor tissues would correlate with patient morbidity and mortality. A clinically annotated tissue microarray (TMA) of primary, recurrent, and metastatic (to lymph node) canine dermal and subcutaneous MCTs was created. Some dogs whose MCTs were included in the TMA did not receive adjunctive treatment after surgical excision of the MCT, whereas others were treated with one or a combination of chemotherapy, radiation, or oral toceranib. Immunohistochemistry for beclin-1, an autophagy protein, was performed followed by digital image analysis. Beclin-1 immunolabeling was higher in recurrent tumors (mean H-score 110.8) than primary MCTs (mean H-score 73.5), and highest in lymph node metastases (mean H-score 138.5) with a significant difference in means (P < .001). While beclin-1 level was not prognostic, it was strongly predictive for survival after adjunctive treatment; dogs with high beclin-1-expressing tumors showed poorer survival compared to those with low beclin-1-expressing tumors (HR = 5.7, P = .02), especially in Kiupel high-grade tumors (HR = 16.3, P = .01). Beclin-1 immunolabeling was the only significant predictive factor by multivariable analysis (P = .04). These findings may improve our ability to predict the response to adjunctive therapy. Importantly, these data suggest that autophagy inhibitors may be useful in improving response to treatment for dogs with high-grade MCTs.
| S-EPMC8679166 | biostudies-literature
Project description:BackgroundMutation analysis of proto-oncogene c-kit (c-kit) is advisable before starting treatment with tyrosine kinase inhibitors in dogs with mast cell tumor (MCT), including those with metastatic disease. Testing is usually performed on primary tumors, assuming that c-kit mutation status does not change in metastasis.Hypothesis/objectivesTo give an insight into the mutational processes and to make a recommendation on the use of c-kit mutational analysis in the clinical setting.AnimalsTwenty-one client-owned dogs with metastatic MCT.MethodsDogs undergoing resection or biopsy for both primary and matched metastatic MCT were prospectively enrolled. Total RNA or DNA was extracted from primary MCT and corresponding metastases. Exons 8, 9, and 11 were amplified by PCR and sequenced. Genetic features between primary MCT and metastases were compared. Their correlation with clinicopathologic features was investigated.ResultsConcordance (mutated or wild-type) of mutational status, evaluable in 21 primary and matched metastatic (20 nodal and 1 splenic) MCTs, was 100%. Three new c-kit mutations were identified. No significant correlation was detected between c-kit mutation and clinicopathologic features.Conclusions and clinical importanceProto-oncogene c-kit mutational status is conserved between any primary and its matched secondary tumor, suggesting that both can be used for c-kit mutational testing. Targeted therapies might be also used to treat metastatic disease.
| S-EPMC4858004 | biostudies-literature
Project description:Mast cell tumors (MCTs) are one of the most common cutaneous malignancies in dogs. Previous studies have reported expression of mast cell-specific proteases chymase and tryptase in canine cutaneous MCTs and in connective tissue and mucosal mast cells. In humans and rodents, mast cells express an additional specific protease, carboxypeptidase A3 (CPA3). In this article, we describe CPA3 immunoreactivity in connective tissue, visceral, mucosal, and neoplastic mast cells in dogs. Positive immunolabeling for CPA3 was observed in nonneoplastic mast cells in 20/20 formalin-fixed paraffin-embedded normal tissues (skin, liver, spleen, intestine), and in 63/63 MCTs irrespective of their histological grade. CPA3 protein expression was comparable to that of c-kit in both the nonneoplastic and neoplastic mast cells. Three distinct labeling patterns (membranous, diffuse, and focal cytoplasmic) were observed for CPA3 in MCTs. The focal cytoplasmic labeling pattern was associated with high-grade MCTs staged with the Kiupel 2-tier grading criteria. We propose CPA3 as a novel immunohistochemical marker for canine mast cells in health and disease.
| S-EPMC8928232 | biostudies-literature
Project description:Mast cell tumours (MCTs) are the most frequent malignant skin neoplasm in dogs. Due to the difficulty in purifying large numbers of canine neoplastic mast cells, relatively little is known about their properties. A reproducible in vitro model is needed to increase the understanding about the phenotype and functional properties of neoplastic mast cells. In the present study, we describe the establishment of primary cocultures of neoplastic mast cells from canine cutaneous MCTs and cancer-associated fibroblasts. We confirmed the inability of canine neoplastic mast cells to remain viable for long periods in vitro without the addition of growth factors or in vivo passages in mice. Using a transwell system, we observed that mast cell viability was significantly higher when there is cell-to-cell contact in comparison to non-physical contact conditions and that mast cell viability was significantly higher in high-grade than in low-grade derived primary cultures. Moreover, the use of conditioned medium from co-cultured cells led to a significantly higher tumoral mast cell viability when in monoculture. Signalling mechanisms involved in these interactions might be attractive therapeutic targets to block canine MCT progression and deserve more in-depth investigations.
| S-EPMC8668961 | biostudies-literature
Project description:Quantitative morphologic parameters assessed in cytologic samples of canine cutaneous mast cell tumors (ccMCTs) may assist with surgical planning and prognostication. Robust cutoffs can be defined, with high reproducibility, for parameters such as the nuclear area (NA). The NA may be determined by morphometry (image analysis, NAI) or by stereology, such as the 2D-nucleator method (NAN); stereologic techniques have not been applied to cytologic specimens of ccMCT, to our knowledge. We retrospectively selected routine cytology smears from 51 ccMCT cases and screened them to determine the percentage of neoplastic mast cells with indistinct nuclear borders; this was repeated after the slides were restained with H&E. The NAI and the NAN were estimated in 100 mast cells per animal in H&E-stained slides. All nuclei were visible in H&E smears, and unbiased quantification was feasible. The NAN was similar to NAI, but less time-consuming. Both the NAN and NAI determined by cytology differed in histologic low- and high-grade ccMCTs, and in histologic grade I plus II versus grade III ccMCTs. Stereologic parameters such as the NAN could be considered as complementary techniques for the cytologic evaluation of ccMCTs.
| S-EPMC8921802 | biostudies-literature
Project description:BACKGROUND:The WW domain-containing oxidoreductase (WWOX) tumor suppressor gene is frequently lost in a variety of solid and hematopoietic malignancies in humans. Dysregulation of WWOX has been implicated as playing a key role in tumor cell survival, DNA damage repair, and genomic stability. The purpose of this study was to characterize WWOX expression in spontaneous canine mast cell tumors (MCTs) and malignant cell lines and investigate the potential contribution of WWOX loss on malignant mast cell behavior. METHODS/RESULTS:WWOX expression is decreased in primary canine MCTs and malignant mast cell lines compared to normal canine bone marrow-cultured mast cells. In transformed canine mastocytoma cell lines, overexpression of WWOX or WWOX knockdown had no effect on mast cell viability. Inhibition of WWOX enhanced clonogenic survival following treatment with ionizing radiation in the C2 mast cell line. Lastly, immunohistochemistry for WWOX was performed using a canine MCT tissue microarray, demonstrating that WWOX staining intensity and percent of cells staining for WWOX is decreased in high-grade MCTs compared to low-grade MCTs. CONCLUSIONS:These data suggest that WWOX expression is attenuated or lost in primary canine MCTs and malignant mast cell lines. Given the observed increase in clonogenic survival in WWOX-deficient C2 mast cells treated with ionizing radiation, further investigation of WWOX and its role in mediating the DNA damage response in malignant mast cells is warranted.
| S-EPMC7603737 | biostudies-literature
Project description:In the present study, canine mast cell tumor (MCT) transcriptome was characterized in order to identify a set of candidate genes potentially useful for MCT classification and prognosis prediction. Fifty-one canine MCT biopsies were enrolled in the study. Isolated and purified total RNAs were individually hybridized to the Agilent Canine V2 4x44k DNA microarray. The comparison of reference differentiated and undifferentiated MCT transcriptome revealed a total of 597 differentially expressed genes (450 up-regulated and 147 down-regulated). The functional analysis of this set of genes provided evidence that they were mainly involved in cell cycle, DNA replication, p53 signaling pathway, nucleotide excision repair and pyrimidine metabolism. Class prediction analysis identified 13 transcripts providing the greatest accuracy of class prediction and divided samples into two categories (differentiated and undifferentiated MCTs). The molecular classification here used was prognostically significant, as the groups were directly correlated with survival time (p = 0.0026). In this study, we analyzed the gene expression profiles of 51 mast cell tumour samples using Agilent Canine V2 4x44k (G2519F, Design ID 021193) DNA microarray platform (60 arrays, 8 replicates) based on single-colour detection (Cyanine-3 only). Microarrays are scanned with Agilent scanner G2565BA (barcode on the left, DNA on the back surface, scanned through the glass) at a resolution of 5 microns; all slides are scanned twice at two different sensitivity settings (XDRHi 100% and XDRLo 10%); the scanner software creates a unique ID for each pair of XDR scans and saves it to both scan image files. Feature Extraction 9.5 uses XDR ID to link the pairs of scans together automatically when extracting data. The signal left, after all the FE processing steps have been completed, is ProcessedSignal that contains the Multiplicatively Detrended, Background-Subtracted Signal.
2014-05-06 | E-GEOD-50433 | biostudies-arrayexpress
Project description:Mast cell tumor (MCT) is a common skin cancer in dogs that has a wide range of clinical behaviors. The purpose of this study was to develop a novel multicolor flow cytometry (FC) panel that will enable the quantification of candidate prognostic markers (Ki-67 and pKIT) in fine needle aspirate (FNA) samples prior to surgical removal of the tumors. FNA of canine MCTs and the NI-1 cell line were utilized to develop a FC panel that includes a viability dye (FVS620, BD Biosciences; 7-AAD, Invitrogen) and the following primary conjugated antibodies: CD117-PE (ACK45, BD Biosciences), pKIT-A647 (polyclonal bs-3242R, BIOSS) and Ki-67-FITC (20Raj1, eBioscience; MIB-1, DAKO). A total of nine FNA samples of canine MCTs were collected, seven out which produced sufficient cells for FC analysis. The Ki-67 antibody clone 20Raj1 produced a positive signal when applied to blood leukocytes but failed to provide robust labeling of neoplastic mast cells. The Ki-67 antibody clone MIB-1 delivered a superior staining quality in both the NI-1 cells and primary MCT cells. CD117-PE signal was adequate post fixation and permeabilization and in the combination of 7-AAD. pKIT produced non-specific staining and was not suitable for this multicolor FC panel. In conclusion, FNA samples of canine MCTs can often yield adequate cell numbers for FC analysis, and a multicolor FC panel was developed that can detect Ki-67 in canine mast cells. This would permit further studies into the potential use of this panel for canine cutaneous and subcutaneous MCT prognostication purposes.
| S-EPMC10704158 | biostudies-literature
Project description:In the present study, canine mast cell tumor (MCT) transcriptome was characterized in order to identify a set of candidate genes potentially useful for MCT classification and prognosis prediction. Fifty-one canine MCT biopsies were enrolled in the study. Isolated and purified total RNAs were individually hybridized to the Agilent Canine V2 4x44k DNA microarray. The comparison of reference differentiated and undifferentiated MCT transcriptome revealed a total of 597 differentially expressed genes (450 up-regulated and 147 down-regulated). The functional analysis of this set of genes provided evidence that they were mainly involved in cell cycle, DNA replication, p53 signaling pathway, nucleotide excision repair and pyrimidine metabolism. Class prediction analysis identified 13 transcripts providing the greatest accuracy of class prediction and divided samples into two categories (differentiated and undifferentiated MCTs). The molecular classification here used was prognostically significant, as the groups were directly correlated with survival time (p = 0.0026).
2014-05-06 | GSE50433 | GEO