Evaluation of expression and function of vascular endothelial growth factor receptor 2, platelet derived growth factor receptors-alpha and -beta, KIT, and RET in canine apocrine gland anal sac adenocarcinoma and thyroid carcinoma

Abstract

Background: Toceranib phosphate (Palladia) has a reported objective response rate of 25% in both canine apocrine gland anal sac adenocarcinoma (AGASACA) and thyroid carcinoma (TC), with stable disease occurring in an additional 50-60% of dogs. The basis for the observed responses to toceranib is not known. The purpose of this study was to evaluate AGASACA and TC samples for the expression and activation of VEGFR2, PDGFRα, PDGFRβ, KIT and RET to assess whether dysregulation of these receptor tyrosine kinases (RTKs) may contribute to the biologic activity of toceranib.

Results: mRNA for VEGFR2, PDGFRα/β, KIT and RET was detected in all AGASACA samples. mRNA for VEGFR2, PDGFRα/β, and KIT was detected in all TC samples, while mRNA for RET was amplified in 10/15 samples. No phosphorylation of VEGFR2, PDGFRα/β, or KIT was observed on the arrays. However, phosphorylation of RET was detected in 54% of the primary AGASACA and 20% of TC. VEGFR2 was expressed in 19/24 primary and 6/10 metastatic AGASACA and 6/15 TC samples. KIT was present in 8/24 primary and 3/10 metastatic AGASACA and 9/15 TC samples. PDGFRα expression was noted in all tumor samples. In contrast PDGFRβ expression was found in only a few tumor samples but was evident in the stroma of all tumor specimens.

Conclusions: Known targets of toceranib are expressed in both AGASAC and TC. Given the observed expression of VEGFR and PDGFRα/β and phosphorylation of RET, these RTKs merit investigation as to their roles in the biology of AGSACA and TC and their contribution to toceranib's activity.

Figure 1

Phospho-RTK array profiling of canine AGASACA and TC tumors. Shown are representative examples of phosphoprotein arrays of paired primary and metastatic AGASACA and TC using the Proteome Profiler Human Phospho-RTK Array Kit. This platform allowed simultaneous screening of 42 different RTKs. Determination of phosphorylation was based on comparison of capture antibody of interest to positive controls located on the periphery of the array. On these sample arrays, positive controls, EGFR, RET, ROR-1, and Tie-1 have been identified for comparison

Figure 2

Immunohistochemistry for VEGFR2, KIT, and PDGFR α/β. The TMA constructed for primary and metastatic AGASACA and TC were probed for VEGFR2, KIT, PDGFRα, and PDGFRβ. Positive immunoreactivity was scored and location of staining was noted. Shown are representative images for primary and metastatic AGASACA and TC for each RTK evaluated