Abstract

RNA is considered as an indicator of the dynamic genetic expression changes in a cell. RNAScope is a commercially available in situ hybridization assay for the detection of RNA in formalin-fixed paraffin-embedded tissue. In this work, we describe the use of RNAScope as a sensitive and specific method for the evaluation of c-KIT messenger RNA (mRNA) in canine mast cell tumor. We investigated the expression of c-KIT mRNA with RNAscope in 60 canine mast cell tumors (MCTs), comparing it with the histological grade and KIT immunohistochemical expression patterns. Our results showed an overall good expression of c-KIT mRNA in neoplastic cells if compared with control probes. We also observed a statistically significant correlation between histological grade and c-KIT mRNA expression. No correlations were found between KIT protein immunohistochemical distribution pattern and c-KIT mRNA expression or histological grade. Our results provide a reference basis to better understand c-KIT mRNA expression in canine MCTs and strongly encourage further studies that may provide useful information about its potential and significant role as a prognostic and predictive biological marker for canine MCTs clinical outcome.

Highlights

  • Canine mast cell tumor (MCT) is one of the most common neoplastic disease in dogs accounting for ∼20% of all canine skin tumors [1, 2]

  • These results confirm that tissue microarray (TMA) constructed from a retrospective formalin fixed paraffin-embedded (FFPE) tissue archive series are fit-for-purpose for the evaluation of c-KIT messenger RNA (mRNA) by RNAScope in canine MCTs; for several reasons, this finding represent a different-making option in biomarker development and discovery

  • We need to address an important critical issue encountered during our experiments in order to offer useful suggestion to optimize the reproducibility of this assay on FFPE canine MCTs

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Summary

Introduction

Canine mast cell tumor (MCT) is one of the most common neoplastic disease in dogs accounting for ∼20% of all canine skin tumors [1, 2]. Cutaneous MCTs can have a very variable biological behavior [1], and several studies have focused on the investigation of predictive factors for MCTs outcome. These factors include the location of the tumor [3, 4], surgical margins [5,6,7,8,9], mitotic activity [10, 11], nuclear morphometry [12, 13], and vascular density [14]. Immunohistochemical expression and protein localization of KIT in neoplastic cells is currently one of the most informative markers for prognostication of canine MCTs [1, 15,16,17]. It has been extensively described that the immunohistochemical localization of KIT protein in neoplastic canine mast cells may have three different patterns: perimembranous labeling (pattern I), focal or stippled cytoplasmic labeling along with loss of perimembranous labeling (pattern II), and diffuse cytoplasmic labeling

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