The objective of the proposed research is to test the capacity to increase tumor cell killing by introducing the herpes simplex virus (HSV) thymidine kinase (TK) gene on a retroviral vector and treating with ganciclovir (GCV). Our approach is to construct a retroviral vector that can be upregulated to express elevated levels of HSV thymidine kinase. We propose to use glucocorticoid induction of known transactivator systems to direct transcription of HSV TK gene from a target promoter element, as well as from the retrovirus LTR. The ability to upregulate HSV TK gene expression will be evaluated in vitro by measuring RNA and protein to determine that transcription, translation, and enzymatic activity can be increased in response to dexamethasone. The effect of increasing thymidine kinase levels on the IC-50 of GCV will be assessed in rat 9L glioma cells in vitro to determine if increased thymidine kinase levels can increase the efficacy of GCV tumor cell killing. The ability to increase the """"""""bystander effect"""""""" will be assessed in vitro in rat 9L glioma cells to determine if higher thymidine kinase levels can increase the number of untransduced neighboring tumor cells that are killed by a tk+ transduced tumor cell.
GTI currently has Phase I and Phase II clinical trials for human brain tumors by injecting retroviral tk producer cells intratumorally and results after GCV treatment are very encouraging. Application of dexamethasone upregulatable retroviral tk vectors to brain tumor gene therapy will not only improve the efficacy of GCV treatment, but also reduce the dosages of GCV and thus toxicity. With these advantages, this upregulatable retroviral tk vector will have good commercial opportunities.
