Assessment of PDGFRβ promoter methylation in canine osteosarcoma using methylation‐sensitive high‐resolution melting analysis

Abstract

Platelet-derived growth factor signalling pathways play a fundamental role in inducing and sustaining the proliferative and prosurvival stimuli in canine osteosarcomas (cOSAs). The increased expression of platelet-derived growth factor receptors (PDGFRs) α and β, and their cognate ligands, were almost invariably observed in cOSAs and OSA-derived cell lines. In particular, overexpression of PDGFRβ-mediated signalling pathways was found in both the tumour microenvironment, where it drives stromal cell recruitment, and in neoangiogenesis, such as in tumour cells where it triggers aberrant proliferation, migration and local invasion. The majority of the pathological consequences of PDGFRβ signalling are because of aberrant expression. In fact, epigenetic dysregulation of oncogenes throughout demethylation of their promoter has emerged as a pivotal mechanism driving oncogenesis. The aim of this study was to assess the methylation status of the PDGFRβ promoter and to clarify its role in modulating the expression of the tyrosine kinase receptor in canine osteosarcoma. The CpG island of the PDGFRβ promoter was identified using a mixed in silico and experimental approach, and a method based upon the methylation-sensitive high-resolution melting assay for quantitatively and precisely assessing the methylation status of the promoter was then set up. The method herein described was then exploited to assess the methylation status of the promoter in a case series of cOSAa. COSAs consistently but variably expressed PDGFRβ. However, the promoter was almost completely demethylated, and its methylation status did not correlate with the expression levels. This finding supported the hypothesis that posttranscriptional regulatory mechanisms may act in cOSAs.

An Update on Molecular Pathways Regulating Vasculogenic Mimicry in Human Osteosarcoma and Their Role in Canine Oncology

“…Meyer et al demonstrated that, in addition to tumor cells, giant cells and osteoblasts in canine OSA were positive for PDGFBB immunostaining, composed of two subunits β, also showing the detection of its mRNA in all study cases ( 109 ). Finally, the dysregulation of the expression levels of PDGFRβ in canine OSA has been attributed to the strong demethylation of CpG sites within the promoter ( 110 ).…”