Analysis of c-KIT mRNA expression and mutations in canine cutaneous mast cell tumours

Introduction: Canine cutaneous mast cell tumour (MCT) is one of the most common neoplasm in dogs. The c-KIT proto-oncogene has been studied due to its involvement in cellular proliferation and MCTs development. In particular c-KIT mutations and aberrant cytoplasmic KIT protein localization had been previously associated with both decreased disease-free and overall survival rates. In the present study, further than protein localization and mutations identification, c-KIT mRNA expression in MCTs compared to non-pathologic tissue samples was evaluated. Materials and Methods: 48 MCT samples and margins were collected during surgical intervention. Immunohistochemistry was performed in formalin-fixed paraffin-embedded samples using the CD117 polyclonal primary antibody. Total RNA was isolated from frozen samples and used for the identification of c-KIT mutations in exons 8, 9, 11 and for the gene expression analysis by quantitative Real Time PCR. Results: For CD117, three staining patterns were recognized: a membrane-associated pattern, a focal cytoplasmic pattern, and a diffuse cytoplasmic pattern. In 4 cases out of 48, c-KIT mutations were identified: 2 internal tandem duplications, 1 deletion and 1 point mutation (S479I). Interestingly, a statistical significant increase of c-KIT mRNA has been observed in grade I and II MCTs towards surgical margins and normal tissue. Conclusions: In the present study, a prevalence of 8% of c-KIT mutations, already evidenced in literature, has been observed in MCTs. Furthermore, the increased c-KIT mRNA in grade I and II MCTs towards surgical margins and normal tissue confirm the importance of c-KIT as an optimal prognostic biomarker in canine MCTs