Comparative study of P-glycoprotein detection by immunohistochemistry and flow cytometry in canine dermal mast cell tumors

Background: Dermal mast cell tumor is one of the most common skin tumors in dogs. Chemotherapy is often needed as treatment. P-glycoprotein is one of the most important transmembrane protein is P-glycoprotein in the development of multidrug resistance (MDR) which can be associated with treatment failure. Objectives: The aim of this study was to evaluate P-gp expression and make comparative examinations with the function of this efflux pump protein by flow cytometry. In addition, we aimed to determine the connection between P-gp results and survival times. Material and Methods: In our study we examined mast cell tumors of 31 dogs. P-gp expression was detected by immunohistochemistry on paraffin embedded tumor tissue samples. We evaluated the percentage, the intensity and the location of the positive reaction (membrane/cytoplasm). P-gp funcion was also determined in 25 patients by flow cytometry analyses. Multidrug resistance Activity Factor (MAF) was determined by the calcein assay. Fluorescent signal intensity was determined by flow cytometry in cells with or without verapamil inhibition. Results and Discussion: According to our examinations, changes of P-gp expression and function (quantifiable with MAF) are positively correlated (r = 0.542, p < 0.05). This observation was true in chemotherapy-treated patients (n = 15), as well. This observation was true in chemotherapy-treated patients (n = 15), too. The P-gp expression and MAF increased with "grade" and the survival times and "grade" had a negative correlation. Interestingly, P-gp expression and MAF had correlated positively with overall survival time and relapse free period. We determined several cut off values to make accurate survival analysis and only with 0.17 MAF cut off showed a negative correlation between MAF and relapse free period (sensitivity: 66.7%; specificity: 71.4%; p = 0.8214). It should be noted that the overall survival times were significantly influenced by several factors, such as cause of death, degree of malignancy, stage, treatments, so they can not be traced back to exclusively one membrane transporter function.