Adenoviral-Mediated Gene Therapy for Thyroid Carcinoma Using Thymidine Kinase Controlled by Thyroglobulin Promoter Demonstrates High Specificity and Low Toxicity

Abstract

A replication defective adenovirus transducing thymidine kinase (TK) gene under the control of the rat thyroglobulin (rTg) promoter (AdrTgtk) was developed to evaluate its cell-specific killing activity in gene therapy. We also developed adenoviruses containing the TK gene driven by the cytomegalovirus (CMV) promoter (AdCMVtk), and luciferase (Luc) gene driven by the rTg or CMV promoter (AdrTgLuc or AdCMVLuc). Luc activity in FRTL-5, HepG2, COS1, rMTC, hMTC, Hela, GH3, T98G, and CA77 cells was measured after infection with AdrTgLuc or AdCMVLuc. FRTL-5 cells produce thyroglobulin (Tg), whereas all other cells are non-Tg-producing cell lines. Transduction by AdCMVLuc caused high Luc activity in all cell lines. However, infection with AdrTgLuc induced Luc activity only in FRTL-5 cells. AdCMVtk or AdrTgtk was used to transduce various cell lines to evaluate the different killing effect. After infection with AdCMVtk vector followed by ganciclovir (GCV) treatment, cell growth was strongly suppressed in all cell lines compared both to noninfected cells and to cells infected by AdCMVLuc in the presence of GCV. When FRTL-5 cells were infected with AdrTgtk followed by GCV treatment, more than 90% were killed, but only a minimal effect was observed in other cell lines, indicating that the Tg promoter transduced TK expression only in Tg-producing cells. When adenovirus is given intravenously, liver and spleen are the major organs infected. A high Luc activity was found in liver and spleen of AdCMVLuc treated animals. No Luc activity was found in liver and spleen of AdrTgLuc-treated animals, indicating that rTg does not transduce Luc expression in non-Tg-producing tissues in vivo. No significant changes of the serum transaminase levels and histologic abnormalities were found in animals treated with AdrTgtk/GCV compared with control animals. High levels of serum transaminases, lymphocyte infiltration, some Kupffer's cell prominence, and extensive single cell hypatocyte death were found in AdCMVtk/GCVtreated animals, indicating severe liver damage induced, as expected, by a noncell-specific promoter. These results indicate that transfer of TK gene driven by the rTg promoter has thyroid cell-specific killing ability in the presence of GCV, little in vivo toxicity, and should be useful in the future for treating thyroid Tg-producing cancers.