Burrai, Giovanni Pietro (2014) Identificazione di nuovi marcatori tumorali nelle neoplasie mammarie del cane: applicazione di tecnologie molecolari innovative. Doctoral Thesis.
Canine mammary tumors (CMT) are the most common neoplasms in female dogs and the discovery of diagnostic, prognostic, and therapeutic markers is essential for improving the disease outcomes and the animal welfare. The aims of this study were to investigate the role of the Receptor tyrosine-kinase ErbB-2 (HER2) and to discover new potential tumor markers in CMT using innovative proteomic approaches. HER2 protein and RNA expressions were determined by immunohistochemistry (IHC) with an antibody extensively used in veterinary medicine and by real-time quantitative RT-PCR (qRT-PCR) as well as by Western Immunoblotting (WB) and Reverse-Phase Protein Arrays (RPPA). An orthogonal validation of HER2 protein expression was carried out using high-resolution mass spectrometry (MS). A diffuse cytoplasmic staining related to antibody lack of specificity was observed by IHC and further confirmed by WB. No differences between benign and malignant neoplasms were noted at mRNA level and the MS technique failed to detect HER2 peptides in the CMT. These results indicate a minor role of HER2 in CMT. Further, 40 proteins were found to be differentially expressed in normal mammary gland and in CMT by MS. Proteins related to glucose and to mitochondrial function as Transketolase, Transketolase like 1, and Prohibitin 2 were also investigate by IHC. The differences observed in hyperplasic and neoplastic lesions compared to normal mammary gland suggest these proteins as new potential markers for CMT.
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