Ca. Richards et Be. Huber, GENERATION OF A TRANSGENIC MODEL FOR RETROVIRUS-MEDIATED GENE-THERAPYFOR HEPATOCELLULAR-CARCINOMA IS THWARTED BY THE LACK OF TRANSGENE EXPRESSION, Human gene therapy, 4(2), 1993, pp. 143-150
Transgenic mice have been generated to determine the tissue-specific e
xpression, safety, and efficacy of a novel chimeric gene that is being
investigated as a test system for virus-directed enzyme prodrug thera
py (VDEPT). The chimeric gene consists of the transcriptional regulato
ry sequences of the albumin gene and the protein-coding sequence of th
e varicella-zoster virus thymidine kinase (VZV-TK) gene inserted into
a retroviral vector. Eight founders were obtained from microinjection
of a nearly full-length proviral fragment containing the chimeric gene
. Liver extracts of the founders and 12 G1 mice were analyzed by enzym
atic and Western blot analysis for the presence of VZV-TK. No VZV-TK e
nzymatic activity or protein was detected. Methylation analysis indica
ted that both the chimeric gene and retroviral sequences were methylat
ed. Treatment of newborn mice with 5-azacytidine or backcrossing into
a DBA/2 genetic background did not result in detectable VZV-TK express
ion or a change in transgene methylation. The poor transgene expressio
n reported here appears to reflect an inherent, continuing problem of
transgenic technology with transgenes that are essentially intact retr
oviral shuttle vectors. These methylation and expression problems are
generally applicable to other animal models for retroviral-mediated ge
ne therapy and should be of interest to researchers as they design and
evaluate preclinical safety and efficacy studies.