In vivo manipulation of interleukin-2 expression by a retroviral tetracycline (tet)-regulated system

Abstract

We have used the tetracycline (tet)-regulated system as described previously to evaluate the applicability of controlled gene expression in cancer gene therapy. As a model gene, we used the human interleukin-2 (IL-2) gene, which has been placed under the transcriptional control of the tetO/promoter. Human melanoma cells were transduced by two modified retroviral tet vectors containing the transactivator regulatory unit and the IL-2 gene driven by the tetO/promoter, respectively. In the absence of tet, IL-2 expression in the target cells was stable over several months. IL-2 production was in the range of 40 U/10 6 cells/24 hours. A fine tuning of IL-2 expression could be achieved by culturing the transduced cells with increasing doses of tet, whereby a concentration of 500 ng/mL tet in the culture medium abrogated IL-2 expression. Most importantly for clinical application, IL-2 expression by the transduced melanoma cells could also be regulated in vivo. When nu/nu mice were inoculated with the transduced tumor cells, they failed to develop tumors. Instead, the inhibition of IL-2 expression in the transduced tumor cells by oral administration of tet led to subcutaneous tumor growth; this growth rate was comparable with the growth rate of subcutaneously inoculated untransduced parental cells. The finding demonstrates the applicability of the tet-regulated system in cancer gene therapy.

Liver-specific expression of interferon γ following adenoviral gene transfer controls hepatitis B virus replication in mice

“…For example, retrovirus-mediated regulated IL-2 expression suppressed growth of melanoma cells in mice. 45 A Dox-regulated adenovirus vector expressing the cytokine interleukin-12 46 was used to treat prostate cancer. A helper dependent vector expressing IFNa in a regulated fashion was used to prevent infection with mouse hepatitis virus.…”

Transgene amplification and persistence after delivery of retroviral vector and packaging functions with E1/E4-deleted adenoviruses

“…Integration of the tk gene was demonstrated by PCR U3/TK [23][24][25] in all tumors derived from H460 cells infected with AdTK/ENV and AdGAG/POL (Fig 8, lanes G-J), whereas as expected, there was no evidence of TK provirus in control tumors derived from AdTK/ENVinfected cells or noninfected cells (Fig 8, lanes A-F). Proviral integration also was confirmed by Southern blot analysis of genomic DNA (XbaI restricted) with a radiolabeled TK probe (data not shown).…”

“…Retrovirus-mediated integration was therefore assessed by PCR analysis of genomic DNA [23][24][25] with a set of primers (U3/TK) specific to the TK provirus, leading to …”

Transcriptional Targeting in Cancer Gene Therapy

“…A tc inducible promoter has been used to drive expression of HSV- tk in breast carcinoma cells and increased the IC50 for GCV by 50 fold, although bystander effects were not enhanced [239]. Both IL-2 and IL-1 β converting enzyme (ICE) have been shown to be tightly regulated, respectively, in human melanoma cells [240] and in a rat brain tumour model [241] in vivo using a similar approach. However, the real beauty of this system has been to selectively regulate transgene expression in certain tissues by combining with tissue/tumour-specific promoters.…”