Molecular cloning of canine Wilms’ tumor 1 for immunohistochemical analysis in canine tissues

Abstract

Wilms’ tumor 1 (WT1) expression has been investigated in various human cancers as a target molecule for cancer immunotherapy. However, few studies have focused on WT1 expression in dogs. Firstly, cDNA of canine WT1 (cWT1) was molecularly cloned from normal canine kidney. The cross-reactivity of the anti-human WT1 monoclonal antibody (6F-H2) with cWT1 was confirmed via Western blotting using cells overexpressing cWT1. Immunohistochemical staining revealed that cWT1 expression was detected in all canine lymphoma tissues and in some normal canine tissues, including the kidney and lymph node. cWT1 is a potential immunotherapy target against canine cancers.

The cell tube block technique and an immunohistochemistry panel including Wilms tumor 1 to assist in diagnosing cavitary effusions in dogs and cats

“…In our study, we observed a lack of staining in lymphoma CTBs, whereas Sakai et al recently reported positivity in 22 cases of canine lymphoma, using the same monoclonal antibody. This controversy could be due to the cytoplasmic vs nuclear WT1 staining patterns.…”

“…Differences between WT1 polyclonal and monoclonal antibodies are well known, but recently immunostains performed with polyclonal antibodies did not correlate with WT1 gene expression in human tissues . Apparently, this problem also occurs in dogs, since multiple nonspecific bands were obtained in the Western blots with the WT1 polyclonal antibodies, which contrasted with a single 50‐55 kDa band (the expected molecular weight of WT1 protein) obtained with the monoclonal antibody used in this study …”

“…WT1 is a tumor suppressor gene, located on chromosome 18 in the dog . Unlike other tumor suppressor genes, WT1 appears to be expressed in selected mesoderm‐derived cell types, such as the mesothelium, kidney, and gonads . In humans, WT1 is considered a useful mesothelial marker, with positive rates of 100% for reactive mesothelium and between 72% and 100% of mesotheliomas .…”

“…It was recently shown that WT1 exists in dogs with a 99% homology to the human WT1 peptide . Using transfection, Western blot, and IHC techniques, it was demonstrated that the human WT1 monoclonal antibody used recognizes canine WT1 .…”

“…It was recently shown that WT1 exists in dogs with a 99% homology to the human WT1 peptide . Using transfection, Western blot, and IHC techniques, it was demonstrated that the human WT1 monoclonal antibody used recognizes canine WT1 . In this vein, for the validation study, we performed IHC for WT1 in a case series (n = 22) of histologically‐confirmed cases of: (a) normal pleura (n = 2); (b) mesothelial hyperplasia (n = 4); (c) lung tumors (n = 5, three primary lung papillary carcinomas and two mammary carcinomatous metastases); (d) mammary tumors (n = 3, two solid and one tubulopapillary carcinoma); (e) ovarian tumors (n = 2, one papillary carcinoma and one granulosa cell tumor); (f) malignant mesotheliomas (n = 6, five epithelioid and one biphasic).…”